Assay with textured surface

ABSTRACT

Among other thing, the present invention provides solution to the problem, particularly, the present invention certain surfaces and certain sample holder to improve the sensitivity, speed, and easy-to-use of an optical signal based assays, such as colorimetric assays or fluorescence assays.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. patent Application Ser. No.16/485,347, filed on Aug. 12, 2019, which is a § 371 national stageapplication of International Application PCT/US2018/018521 filed on Feb.16, 2018, which claims the benefit of priority to U.S. ProvisionalApplication (“U.S. Patent Application” hereafter) No. 62/460,088, filedon Feb. 16, 2017, U.S. Patent Application No. 62/460,091, filed on Feb.16, 2017, U.S. Patent Application No. 62/460,083, filed on Feb. 16,2017, U.S. Patent Application No. 62/460,076, filed on Feb. 16, 2017,U.S. Patent Application No. 62/460,075, filed on Feb. 16, 2017, U.S.Patent Application No. 62/460,069, filed on Feb. 16, 2017, U.S. PatentApplication No. 62/460,062, filed on Feb. 16, 2017, U.S. PatentApplication No. 62/460,047, filed on Feb. 16, 2017, U.S. PatentApplication No. 62/459,972, filed on Feb. 16, 2017, U.S. PatentApplication No. 62/459,920, filed on Feb. 16, 2017, PCT Application No.PCT/US18/18405, filed on Feb. 15, 2018, PCT Application No.PCT/US18/18108, filed on Feb. 14, 2018, PCT Application No.PCT/US18/18007, filed on Feb. 13, 2018, PCT Application No.PCT/US18/17716, filed on Feb. 9, 2018, PCT Application No.PCT/US18/17713, filed on Feb. 9, 2018, PCT Application No.PCT/US18/17712, filed on Feb. 9, 2018, PCT Application No.PCT/US18/17504, filed on Feb. 8, 2018. PCT Application No.PCT/US18/17501, filed on Feb. 8, 2018. PCT Application No.PCT/US18/17499, filed on Feb. 8, 2018. PCT Application No.PCT/US18/17489, filed on Feb. 8, 2018. PCT Application No.PCT/US18/17492, filed on Feb. 8, 2018. PCT Application No.PCT/US18/17494, filed on Feb. 8, 2018. PCT Application No.PCT/US18/17502, filed on Feb. 8, 2018, and PCT Application No.PCT/US18/17307, filed on Feb. 7, 2018, the contents of which are reliedupon and incorporated herein by reference in their entirety. The entiredisclosure of any publication or patent document mentioned herein isentirely incorporated by reference.

FIELD

Among other things, the present invention is related to devices andmethods of performing biological and chemical assays, devices andmethods of performing a biological and chemical using colorimetricapproaches.

BACKGROUND

In bio/chemical assaying, there is a need to enhance light signal from athin sample. For example, in colorimetric assay, when the samplethickness is very thin (e.g. 100 um (micron) or less), the color becomevery faint, become difficult to be observed, limiting the sensitivity ofa colorimetric assay.

SUMMARY OF INVENTION

The following brief summary is not intended to include all features andaspects of the present invention. Among other thing, the presentinvention provides solutions to the to improve the sensitivity, speed,and easy-to-use of assaying by optical signal, such as colorimetricassays or fluorescent assays.

BRIEF DESCRIPTION OF THE DRAWINGS

The skilled artisan will understand that the drawings, described below,are for illustration purposes only. The drawings are not intended tolimit the scope of the present teachings in any way. The drawings arenot entirely in scale. In the figures that present experimental datapoints, the lines that connect the data points are for guiding a viewingof the data only and have no other means.

FIG. 1 -A illustrates an example of opened assembled colorimetric assaysample card comprising a bottom plate, a top plate and an aluminumhinge, in accordance with an embodiment of the present invention.

FIG. 1 -B and FIG. 1 -C illustrate an example of bottom plate of thecolorimetric assay sample card having textured microstructures on topsurface, in accordance with an embodiment of the present invention.

FIG. 1 -D and FIG. 1 -E illustrate an example of top plate of thecolorimetric assay sample card having pillar arrays of uniform heightson bottom surface, in accordance with an embodiment of the presentinvention.

FIG. 1 -F and FIG. 1 -G illustrates an example of ready-to-testcolorimetric assay sample card comprising a bottom plate, a top plate,an aluminum hinge and sample liquid between top and bottom plates, inaccordance with an embodiment of the present invention.

FIG. 2 -A illustrates a test apparatus of colorimetric measurement ofsample with textured surfaces using side illumination of a fiber.

FIG. 2 -B, FIG. 2 -C and FIG. 2 -D illustrates a test apparatus ofcolorimetric measurement of sample with textured surfaces using ringillumination of a fiber.

FIG. 3 is an illustration of a CROF (Compressed Regulated Open Flow)embodiment. Panel (a) illustrates a first plate and a second platewherein the first plate has spacers. Panel (b) illustrates depositing asample on the first plate (shown), or the second plate (not shown), orboth (not shown) at an open configuration. Panel (c) illustrates (i) using the two plates to spread the sample (the sample flow between theplates) and reduce the sample thickness, and (ii) using the spacers andthe plate to regulate the sample thickness at the closed configuration.The inner surface of each plate may have one or a plurality of bindingsites and or storage sites (not shown).

FIG. 4 A diagram of a process of testing heavy metal in water.

DETAILED DESCRIPTION OF EXEMPLARY EMBODIMENTS

The following detailed description illustrates some embodiments of theinvention by way of example and not by way of limitation. The sectionheadings and any subtitles used herein are for organizational purposesonly and are not to be construed as limiting the subject matterdescribed in any way. The contents under a section heading and/orsubtitle are not limited to the section heading and/or subtitle, butapply to the entire description of the present invention.

The citation of any publication is for its disclosure prior to thefiling date and should not be construed as an admission that the presentclaims are not entitled to antedate such publication by virtue of priorinvention. Further, the dates of publication provided can be differentfrom the actual publication dates which can need to be independentlyconfirmed.

A. QMAX Colorimetric Assay with Textures Reflective Scattering Surfaces

In an assay involving a detection of light signal, such as colorimetricassay or fluorescese assay, a small container to hold a liquid sampleand passes a light beam though the sample to measure the light or thecolor of the sample. When the sample is very thin, the light or colorbecomes faint and difficult measure.

The present invention provides, among other thing, solution to get astronger optical signal in a thin sample.

One novelty of the present invention is to use QMAX card (that has twomovable plates) to make a sample into a very uniform thin layer (lessthan 200 um).

Another novelty of the present invention is to use a textured reflectivesurface on a surface of one of the two plates to enhance an opticalsignal, particularly for colorimetric assay and/or fluorescence assay.

In the present invention, we observed that the color signal of acolorimetric assay can be significantly increased by using a reflectivetextured surface as one of the wall of the chamber can significantlyincrease the color signal.

According the present invention, a device uses to plates to sandwich asample into a thin layer, wherein one of the plate is transparent andthe other plate has a textured reflective surface on its sample contactarea. The probing light enters the sample from the transparent plate,goes through the sample, and diffusively reflected by the texturedsurface back to the transparent plate. We have observed that sucharrangement can significantly increase the color signal even the sampleas thin as 30 um or less.

Furthermore, according to the present invention, the device furthercomprise a dry reagent coated on one of the plate, so that a liquidsample can dropped on one or both of the plate, close the plates, andthen measurement. The sample thickness can be 150 um or less, making thedry regent mixed with the sample in a short time, to speed up the totalmeasurement time.

The terms “CROF Card (or card)”, “COF Card”, “QMAX-Card”, “Q-Card”,“CROF device”, “COF device”, “QMAX-device”, “CROF plates”, “COF plates”,and “QMAX-plates” are interchangeable, except that in some embodiments,the COF card does not comprise spacers; and the terms refer to a devicethat comprises a first plate and a second plate that are movablerelative to each other into different configurations (including an openconfiguration and a closed configuration), and that comprises spacers(except some embodiments of the COF) that regulate the spacing betweenthe plates. The term “X-plate” refers to one of the two plates in a CROFcard, wherein the spacers are fixed to this plate. More descriptions ofthe COF Card, CROF Card, and X-plate are described in in PCT Application(designating U.S.) Nos. PCT/US2016/045437 and PCT/US0216/051775, whichwere respectively filed on Aug. 10, 2016 and Sep. 14, 2016, U.S.Provisional Application No. 62/456,065, which was filed on Feb. 7, 2017,U.S. Provisional Application No. 62/456,287, which was filed on Feb. 8,2017, and U.S. Provisional Application No. 62/456,504, which was filedon Feb. 8, 2017, all of which applications are incorporated herein intheir entireties for all purposes.

Device_0 (General)

N1. In some embodiments, according to the present invention, a devicefor assaying a sample using optical signal, comprising:

a first plate, a second plate, spacers, and a textured surface, wherein:

-   -   i. the first and second plates are movable relative to each        other into different configurations;    -   ii. one or both plates are flexible;    -   iii. the second plate has, on its inner surface, have textured        structures for scattering the light illuminated on the surface;    -   iv. the textured surface can be, but is not limited to a bumpy,        wavy roughly surface;    -   v. the textured surface is periodic or aperiodic;    -   vi. the textured surface's average roughness range is preferred        to be, but is not limited to 2 um-5 um;    -   vii. the spacers are fixed to the inner surface of the first        plate and have a predetermined uniform height;    -   viii. the preferred height of spacers is larger than the average        roughness of the textured surface and smaller than 100 um;

wherein on of the configuration is an open configuration, in which: twoplates are partially or entirely separated apart, the spacing betweenthe plates is not regulated by the spacers, and the sample is depositedon one or both of the plates;

wherein on of the configuration is a closed configuration, which isconfigured after the sample deposition in the open configuration, and inthe closed configuration: at least part of the deposited sample iscompressed by the two plates into a continuous layer; wherein the sampleis in liquid form.

-   Device_C1 (for Colorimetric Signal)

A sample handling device for enhancing optical signal (Q-card),comprising:

A first plate, a second plate, spacers and textured surface, wherein:

-   -   i. the plates are movable relative to each other into different        configurations;    -   ii. one or both plates are flexible;    -   iii. the second plate has, on its inner surface, have textured        structures for scattering the light illuminated on the surface;    -   iv. the textured surface can be, but is not limited to a bumpy,        wavy roughly surface;    -   v. the textured surface can be periodic or aperiodic;    -   vi. the textured surface's average roughness range is preferred        to be, but is not limited to 2 um-5 um;    -   vii. the spacers are fixed to the inner surface of the first        plate and have a predetermined uniform height;    -   viii. the preferred height of spacers is larger than the average        roughness of the textured surface and smaller than 100 um;

wherein on of the configuration is an open configuration, in which: twoplates are partially or entirely separated apart, the spacing betweenthe plates is not regulated by the spacers, and the sample is depositedon one or both of the plates;

wherein on of the configuration is a closed configuration, which isconfigured after the sample deposition in the open configuration, and inthe closed configuration: at least part of the deposited sample iscompressed by the two plates into a continuous layer; wherein the sampleis in liquid form.

In some embodiments, the textured surface is made of opaque whitematerial.

-   Device_C2 (for Colorimetric Signal)

A sample handling device for enhancing optical signal (Q-card),comprising:

A first plate, a second plate, spacers and textured surface, wherein:

-   -   i. the plates are movable relative to each other into different        configurations;    -   ii. one or both plates are flexible;    -   iii. the second plate has, on its inner surface, have textured        structures for scattering the light illuminated on the surface;    -   iv. the textured surface can be, but is not limited to a bumpy,        wavy roughly surface;    -   v. the textured surface can be periodic or aperiodic;    -   vi. the textured surface's average roughness range is preferred        to be, but is not limited to 2 um-5 um;    -   vii. the spacers are fixed to the inner surface of the first        plate and have a predetermined uniform height;    -   viii. the preferred height of spacers is larger than the average        roughness of the textured surface and smaller than 100 um;

wherein on of the configuration is an open configuration, in which: twoplates are partially or entirely separated apart, the spacing betweenthe plates is not regulated by the spacers, and the sample is depositedon one or both of the plates;

wherein on of the configuration is a closed configuration, which isconfigured after the sample deposition in the open configuration, and inthe closed configuration: at least part of the deposited sample iscompressed by the two plates into a continuous layer; wherein the sampleis liquid form.

In some embodiments, the textured surface is made of semi-opaque whitematerial, and the transmissivity is 10%˜30%.

-   Device_F (for Fluorescent Signal)

A sample handling device for enhancing optical signal (Q-card),comprising:

A first plate, a second plate, spacers and textured surface, wherein:

-   -   i. the plates are movable relative to each other into different        configurations;    -   ii. one or both plates are flexible;    -   iii. the second plate has, on its inner surface, have textured        structures for scattering the light illuminated on the surface;    -   iv. the textured surface can be, but is not limited to a bumpy,        wavy roughly surface;    -   v. the textured surface can be periodic or aperiodic;    -   vi. the textured surface's average roughness range is preferred        to be, but is not limited to 2 um-5 um;    -   vii. the spacers are fixed to the inner surface of the first        plate and have a predetermined uniform height;    -   viii. the preferred height of spacers is larger than the average        roughness of the textured surface and smaller than 100 um;

wherein on of the configuration is an open configuration, in which: twoplates are partially or entirely separated apart, the spacing betweenthe plates is not regulated by the spacers, and the sample is depositedon one or both of the plates;

wherein on of the configuration is a closed configuration, which isconfigured after the sample deposition in the open configuration, and inthe closed configuration: at least part of the deposited sample iscompressed by the two plates into a continuous layer;

wherein the sample is in liquid form.

In some embodiments, the textured surface is made of opaque whitematerial or coated with reflective metal film, the metal film can be,but is not limited to aluminum, silver and gold. The preferred thicknessrange of the metal film is preferred to be, but not limited to be 10nm˜100 nm.

-   Apparatus (for Colorimetric Signal).A1

A testing apparatus, comprising:

-   -   a) a sample handling device for enhancing optical signal        (Device_C1), according to the above device claim;    -   b) a mobile computing device having a camera module and a light        source;    -   c) an Illumination optics, comprising a tilted optical fiber;    -   d) an external lens;

wherein the light source emits white light;

wherein the light source and camera module are on the same face of themobile computing device;

wherein the Q-card is put right under camera module, the preferreddistance between them is 15 mm-20 mm;

wherein the external lens is put between the Q-card and camera module sothat the sample in Q-card is in the working distance of camera module,and the preferred focal length of external lens is 12˜18 mm, and thedistance between lens and camera module is preferred to be as small aspossible and no larger than 3 mm;

wherein the optical fiber guide the light emitted from the light sourceto illuminate on the sample area right under the camera module;

wherein one end face of the optical fiber is put under the aperture ofthe light source, and the distance between them is preferred to be assmall as possible and no larger than 3 mm;

wherein the diameter of the optical fiber is configured to be equal tothe diameter of the light source aperture;

wherein the tilt angle in which the optical fiber is mounted is set tomake the center light beam emitted out from the fiber illuminate on thesample area right under the camera module.

-   Apparatus (for Colorimetric Signal).A2

A testing apparatus, comprising:

-   -   a) a sample handling device for enhancing optical signal        (Device_C2), according to the above device claim;    -   b) a mobile computing device having a camera module and a light        source;    -   c) an Illumination optics, comprising a pair of reflective        mirrors;    -   d) an external lens;

wherein the light source emits white light;

wherein the light source and camera module are on the same face of themobile computing device;

wherein the Q-card is put right under camera module, the preferreddistance between them is 5˜10 mm;

wherein the external lens is put between the Q-card and camera module sothat the sample in Q-card is in the working distance of camera module,and the preferred focal length of external lens is 4˜8 mm, and thepreferred distance between lens and camera module is preferred to be assmall as possible and no larger than 3 mm;

wherein the illumination optics turns the light emitted from the lightsource to back-illuminate the sample on Q-card, and each mirror turnsthe light by 90 degree;

wherein the mirrors are mounted under the Q-card, and one mirror is in aline with the light source, and another one is in a line with the cameramodule, and the preferred distance between the Q-card and mirrors is 5mm-10 mm.

-   Apparatus (for Colorimetric Signal).A3

A testing apparatus, comprising:

-   -   a) a sample handling device for enhancing optical signal        (DeviceS2), according to the above device claim;    -   b) a mobile computing device having a camera module;    -   c) a separate light source;    -   d) an external lens;

wherein the light source emits white light, and the light source is putunder the Q-card and in a line with the camera module, and the preferreddistance between the light source and Q-card is 5 mm-10 mm.

wherein the Q-card is put right under camera module, the preferreddistance between them is 5˜10 mm;

wherein the external lens is put between the Q-card and camera module sothat the sample in Q-card is in the working distance of camera module,and the preferred focal length of external lens is 4˜8 mm, and thepreferred distance between lens and camera module is preferred to be assmall as possible and no larger than 3 mm;

-   Optical Signals

According the present invention, the optical signal that can by enhancedby the textured surfaces of the device of any prior embodiment, isselected from a group of colors in the sample, fluorescence,luminescence (electrical, chemical, photo, or electrical-chemical),and/or other light from emitters.

-   Apparatus for Fluorescent Signal).A4

A testing apparatus, comprising:

-   -   a) a sample handling device for enhancing optical signal        (Device_F), according to the above device claim;    -   b) a mobile computing device having a camera module;    -   c) a separate light source;    -   d) an Illumination optics, comprising tilted reflective mirror;    -   e) filters, comprising a long pass filter and a short pass        filter;    -   f) an external lens;

wherein the light source is a laser diode;

wherein the tilt mirror turns the light emitted from the light source toilluminate on the sample area right under the camera module;

wherein the light illuminates on the sample in an oblique angle,preferred angle is >60 degree;

wherein the Q-card is put right under camera module, the preferreddistance between them is 15 mm-20 mm;

wherein the external lens is put between the Q-card and camera module sothat the sample in Q-card is in the working distance of camera module,and the preferred focal length of external lens is 12 mm-18 mm, and thepreferred distance between lens and camera module is preferred to be assmall as possible and no larger than 3 mm;

wherein the short pass filter is put in front of the aperture of thelight source;

wherein the long pass filter is put between the external lens and cameramodule.

-   Method

A method for analyzing the optical signal of sample, comprising thesteps of:

-   -   a) collecting a sample liquid;    -   b) obtaining a device of any prior embodiment;    -   c) depositing the sample on one or both of the plates of the        device when the plates are in an open configuration;    -   d) bringing the two plates together and pressing the plates into        the closed configuration so that the sample forms a liquid layer        between the two plates;    -   e) inserting the device into the testing apparatus;    -   f) turning on the light source of the testing apparatus;    -   g) using camera module capture an image of the sample; and    -   h) the mobile computing device process the image to analyze        colorimetric or fluorescent signal of the image to get some        property of the sample.

-   Application

The Q-card device, testing apparatus and the method above can be appliedto detect presence and level of the analyte of interest in the followingfields:

-   -   1) Food science and safety: testing pH, ammonia, nitrite,        nitrate, heavy metal, bacteria level, etc. in drinking water;        testing bacteria, lactose, additive, particular protein level,        etc. in milk;    -   2) Personal health monitoring: glucose, alcohol, etc. in saliva,        urine and breath.    -   3)

In some embodiments, a device for enhancing an optical signal inassaying comprises:

a first plate, a second plate, spacers, and a light scattering layer,wherein:

-   -   i. the first and second plates are movable relative to each        other into different configurations, and have, on its respective        inner surface, a sample contact area for contacting a sample        that contains an analyte;    -   ii. one or both of the plates are flexible;    -   iii. the first plate is transparent to the light, and    -   iv. the second plate substantially reflect light and comprises        an inner surface a light scattering layer that has a rough        topology;

wherein one of the configurations is an open configuration, in which theaverage spacing between the inner surfaces of the two plates is at least200 um, and the sample is deposited on one or both of the plates;

wherein another of the configurations is a close configuration, which isconfigured after the sample deposition in the open configuration, and inthe closed configuration: at least part of the sample is between the twoplates and the average spacing between the inner surfaces of the platesis less than 200 um; and

wherein in the closed configuration, the light scattering layer enhancestrapping a probe light between the inner surface of the two plates.

In some embodiments, in the device, the light scattering surface of thesecond plate comprises:

-   -   i. the textured surface can be, but is not limited to a bumpy,        wavy roughly surface;    -   ii. the textured surface can be periodic or aperiodic;    -   iii. the textured surface's average roughness range is preferred        to be, but is not limited to 2 um-5 um; or    -   iv. the spacers are fixed to the inner surface of the first        plate and have a predetermined uniform height; and    -   v. a combination of thereof.

-   C1. The device or system of any prior embodiments, wherein the light    scattering layer can be made of highly reflectively opaque white    material with reflectivity at least 50%, 60%, 70%, 80%, 90%, 100%,    or in a range between any of the two values.

-   C2. The device or system of any prior embodiments, wherein the    reflection spectrum of the light scattering surface is within the    range of 300 nm to 1000 nm.

-   C3. The device or system of any prior embodiments, wherein the light    scattering layer can be made of semi-opaque white material, and the    transmissivity is 10%˜30%.

-   C4. The device or system of any prior embodiments, wherein the light    scattering layer can be made of reflective metal film, wherein the    light scattering layer can be made of opaque white dielectric film.

-   C5. The device or system of any prior embodiments, wherein the the    light scattering layer has textured surfaces with R_(a) (arithmetic    average roughness) of 0.5 um-200 um, R_(sm) (mean spacing of the    asperities) of >0.5 um and RΔ_(a) (average slop of the profile) >0.1

-   C6. The device or system of any prior embodiments, wherein the    textured surface can be periodic or aperiodic, wherein the shape of    a single feature on the textured surface can be but not limited to    square, triangle, sharp corner.

-   C7. The device or system of any prior embodiments, wherein the    height of spacers is larger than the average roughness of the    textured surface and smaller than 200 um.

The device or system of any prior embodiments, the average roughnessheight (R_(a)) of the textured reflective need to be at least 20% of thewavelength of the illumination light and can be up to 5-fold of thespacing between the first plate and second plate, or in range betweenthese two values;

The device or system of any prior embodiments, the average lateralfeature size (b_(a)) need to be at least 20% and up to 10-fold of thewavelength of the illumination light, or in range between these twovalues;

The device or system of any prior embodiments, the average period(b_(a)) need to be at least 50% and up to 1000-fold of the wavelength ofthe illumination light, or in range between these two value.

FIG. 1 -A is the schematic illustration of colorimetric assay samplecard 1 in open status. Sample card 1 comprises a top plate 12, a bottomplate 11 and an aluminum hinge 13. Hinge 13 attach top plate 12 tobottom plate 13.

The height of the random scattering structures is from 1 nm to 200 nm,from 1 nm to 300 nm, and from 1 nm to 5000 nm.

In some embodiments, the reflection surface can be done by randomnanoparticles of the same size or different size.

In some embodiments, the reflective range from 50% to 100%, from 30% to100% and from 50% to 80%. They are either wide band or narrow band inspectrum,

FIG. 1 -B and FIG. 1 -C are the schematic illustrations of bottom plate11 in sample card 1, shown from isometric view and cross-section viewrespectively. The material for the bottom plate 11 is nonabsorbent andhas opaque white color. It can be, but is not limited to, whitepolyethylene. The bottom plate 11 has textured surface 11S on one of itstop surface (i.e. the surface facing the top plate 12). The texturedsurface 11S can be random microstructures or periodic microstructures.For random microstructures, it can be, but is not limited to, a bumpy,wavy or rough surface. In one embodiment, the textured surface is thebumpy surface of the matte finish of the white polystyrene sheet withaverage roughness of 2˜3 um. For periodic microstructures, it can be,but is not limited to round, rectangular and triangular pillarsprotruding from a surface of the bottom plate with a square, hexagonalor other lattices. A notch 11N is fabricated on one side of bottom plate11 to make it easy to open top plate 12. A triangle gap 11C isfabricated at one corner of bottom plate 11 to easily differentiate thefront and bottom surface of bottom plate 11.

FIG. 1 -D and FIG. 1 -E are the schematic illustrations of top plate 12in sample card 1, shown from isometric view and cross-section viewrespectively. The material for the top plate is transparent and can be,but is not limited to, PMMA. On bottom surface of the top plate (i.e.the surface facing the bottom plate 11), there are periodic micro-sizepillar arrays 12S with uniform heights. The pillar array can be, but isnot limited to, rectangular pillars with square lattice. In oneembodiment, the top plate is made of PMMA of 175 um thickness and thepillar array has a square lattice with the period of 120 um*110 um. Andeach pillar has the rectangular shape with the dimension of 30 um*40 umand the pillar height is 30 um. A triangle gap 12C is fabricated at onecorner of bottom plate 12 to easily differentiate the front and bottomsurfaces of top plate 12.

FIG. 1 -F and FIG. 1 -G are the schematic illustrations of colorimetricassay sample card 1 in closed status with sample liquid, shown fromisometric view and cross-section view respectively. The sample liquid 1Lis embedded between top plate 12 and bottom plate 11. The texturedsurface 11S of bottom plate 11 is towards the bottom surface of the topplate 12 with pillar array 12S. The average liquid layer thickness ofthe sample liquid 1L is uniform and determined by height of the pillararray 12S on top plate 12. Hence, the volume of the sample liquid 1Lholding in sample card 1 per unit area in this present invention can beaccurately determined. Under the illumination of white light, texturedsurface 11S of bottom plate 11 helps deflect the light beams to increasethe light path inside the sample liquid layer 1L. Hence, lightabsorption by the colored compounds in sample liquid 1L is increased andthe color change is enhanced.

FIGS. 2 -A, 2-B and 2-C are the schematic views showing details ofsystem 10 reading a colorimetric card, and particularly of device 13.FIG. 15 -A is the sectional view showing details of device 13. And FIG.15 -B and FIG. 15 -C are the schematic views only showing theconfiguration of the optics elements in device 13. These figuresillustrate the functionality of the elements that were described abovewith reference to FIG. 14 . The light emitted from light source 1L iscoupled into side-emitting optical fiber ring 135 from the two end facesof fiber ring 135 and travels inside along the ring. Beam B1 is emittedout from the side wall of fiber ring and go through the diffuser film136. Beam B1 illuminates the sample area of colorimetric sample card 138right under the camera 1C from front side to create uniformillumination. The illuminated sample area absorbs part of beam B1 andreflects the beam B1 to beam B2. Beam B2 is collected by lens 133 andgets into camera 1C Lens 133 creates an image of the sample area on theimage sensor plane of camera 1C. Smartphone 1 captures and processes theimage to analyze the color information in the image to quantify thecolor change of the colorimetric assay.

In some embodiments, no spacers are used in regulating the samplethickness between the two plates.

In some embodiments, the textured reflective surface of the plate hasone or a combination of each of the parameters:

-   Optical Signal Enhanced by Textured Surfaces

Colorimetric assay's signal can be enhanced by the textured surfaces. Incolorimetric assay, under the illumination of white light, a specificwavelength of light is absorbed by the colored compounds, which resultsin the color change. Hence, to get stronger color change signal, morelight of the specific absorbing wavelength of the color compounds needsto get absorbed. And based on Beer-Lambert law which determines how muchpercent of light is absorbed when light passing through a lightabsorbing medium, the way to increase the light absorption in acolorimetric assay is to increase the light path in the sample liquid.Compared to a flat reflective surface, the textured surface can make thesmall-angle incident light be reflected to a large-angle emergent lightby scattering to increase the light path in the sample liquid. Andtextured surface can scatter the incident light several times in thesample liquid to increase the light path before the light emits out.

Fluorescent signal of an assay can also be enhanced by the texturedsurface. In fluorescent assay, under the illumination of excitationlight with a specific wavelength, the emitting fluorescent intensity isproportional to the product of fluorescent dye's quantum yield andabsorbed amount of excitation light. The textured surface increases thelight path of excitation light in the sample liquid by scattering hencemore excitation light is absorbed by the fluorescent molecules.

A test apparatus comprises the device, a light source, an optical fiberand an imager

wherein the light source emits light within wavelength range of 300 nmto 1000 nm;

wherein the light source and imager are on a same plane;

wherein the Q-card is put right under the imager, the preferred distancebetween them is 15 mm-20 mm;

wherein the optical fiber guide the light emitted from the light sourceto illuminate on the sample area right under the camera module;

wherein one end face of the optical fiber is put under the aperture ofthe light source, and the distance between them is preferred to be assmall as possible and no larger than 10 mm;

wherein the diameter of the optical fiber is configured to be equal tothe diameter of the light source aperture;

wherein the tilt angle in which the optical fiber is mounted is set tomake the center light beam emitted out from the fiber illuminate on thesample area right under the camera module.

A test apparatus comprises the device, a light source, a ring-shapeoptical fiber and an image,

wherein the light source emits light within wavelength range of 300 nmto 1000 nm;

wherein the ring fiber is a side-emitting optical fiber that canoutcouple light from the wall of the fiber;

wherein the ring fiber is in a circle around the imager;

wherein the Q-card is put right under the imager, the preferred distancebetween them is 15 mm-20 mm;

wherein the light emits from the side of the ring-shape fiber toilluminate the sample;

wherein both end faces of the ring-shape optical fiber are put under theaperture of the light source;

wherein a light diffuser is put between the ring-shape fiber and sampleto diffuse the light emitting from the ring fiber;

-   B. Spacers, Hinges, and Opening Notch

In biological and chemical assaying (i.e. testing), a device and/or amethod that simplifies assaying operation or accelerates assaying speedis often of great value.

In the QMAX (Q: quantification; M: magnifying; A: adding reagents; X:acceleration; also known as compressed regulated open flow (CROF)) assayplatform, a QMAX card uses two plates to manipulate the shape of asample into a thin layer (e.g. by compressing) (as illustrated in FIG. 1). In certain embodiments, the plate manipulation needs to change therelative position (termed: plate configuration) of the two platesseveral times by human hands or other external forces. There is a needto design the QMAX card to make the hand operation easy and fast.

In QMAX assays, one of the plate configurations is an openconfiguration, wherein the two plates are completely or partiallyseparated (the spacing between the plates is not controlled by spacers)and a sample can be deposited. Another configuration is a closedconfiguration, wherein at least part of the sample deposited in the openconfiguration is compressed by the two plates into a layer of highlyuniform thickness, the uniform thickness of the layer is confined by theinner surfaces of the plates and is regulated by the plates and thespacers.

In a QMAX assay operation, an operator often needs to add assay reagentsinto the sample in a controlled fashion. For instance, in someembodiments, the reagents (e.g. detection agent and binding agent) arecoated on the plate surface of the QMAX device, and some reagents (e.g.detection agent) are released into the sample at an appropriate timingduring the assay process. Among many others, in some cases, it isdesirable for the detection agent to be added after the substantialbinding of the target analyte by the binding agent. In other cases, itis desirable to add the detection agent after the formation of the thinfilm of the sample. In other cases, it is desirable to delay theaddition of the detection agent by a specified time period. The presentinvention is to provide devices and methods for achieving these goals aswell as for making bio/chemical sensing (including, not limited to,immunoassay, nucleic assay, electrolyte analysis, etc.) faster, moresensitive, less steps, easy to perform, smaller amount of samplesrequired, less or reduced (or no) needs for professional assistance,and/or lower cost, than many current sensing methods and devices.

The term “compressed open flow (COF)” refers to a method that changesthe shape of a flowable sample deposited on a plate by (i) placing otherplate on top of at least a part of the sample and (ii) then compressingthe sample between the two plates by pushing the two plates towards eachother; wherein the compression reduces a thickness of at least a part ofthe sample and makes the sample flow into open spaces between theplates. The term “compressed regulated open flow” or “CROF” (or“self-calibrated compressed open flow” or “SCOF” or “SCCOF”) (also knownas QMAX) refers to a particular type of COF, wherein the final thicknessof a part or entire sample after the compression is “regulated” byspacers, wherein the spacers are placed between the two plates. Here theCROF device is used interchangeably with the QMAX device.

The term “spacers” or “stoppers” refers to, unless stated otherwise, themechanical objects that set, when being placed between two plates, alimit on the minimum spacing between the two plates that can be reachedwhen compressing the two plates together. Namely, in the compressing,the spacers will stop the relative movement of the two plates to preventthe plate spacing becoming less than a preset (i.e. predetermined)value.

The term “a spacer has a predetermined height” and “spacers have apredetermined inter-spacer distance” means, respectively, that the valueof the spacer height and the inter spacer distance is known prior to aQMAX process. It is not predetermined, if the value of the spacer heightand the inter-spacer distance is not known prior to a QMAX process. Forexample, in the case that beads are sprayed on a plate as spacers, wherebeads are landed at random locations of the plate, the inter-spacerdistance is not predetermined. Another example of not predeterminedinter spacer distance is that the spacers moves during a QMAX processes.

The term “a spacer is fixed on its respective plate” in a QMAX processmeans that the spacer is attached to a location of a plate and theattachment to that location is maintained during a QMAX (i.e. thelocation of the spacer on respective plate does not change) process. Anexample of “a spacer is fixed with its respective plate” is that aspacer is monolithically made of one piece of material of the plate, andthe location of the spacer relative to the plate surface does not changeduring the QMAX process. An example of “a spacer is not fixed with itsrespective plate” is that a spacer is glued to a plate by an adhesive,but during a use of the plate, during the QMAX process, the adhesivecannot hold the spacer at its original location on the plate surface andthe spacer moves away from its original location on the plate surface.

The term “open configuration” of the two plates in a QMAX process meansa configuration in which the two plates are either partially orcompletely separated apart and the spacing between the plates is notregulated by the spacers

The term “closed configuration” of the two plates in a QMAX processmeans a configuration in which the plates are facing each other, thespacers and a relevant volume of the sample are between the plates, therelevant spacing between the plates, and thus the thickness of therelevant volume of the sample, is regulated by the plates and thespacers, wherein the relevant volume is at least a portion of an entirevolume of the sample.

The term “a sample thickness is regulated by the plate and the spacers”in a QMAX process means that for a give condition of the plates, thesample, the spacer, and the plate compressing method, the thickness ofat least a port of the sample at the closed configuration of the platescan be predetermined from the properties of the spacers and the plate.

The term “inner surface” or “sample surface” of a plate in a QMAX devicerefers to the surface of the plate that touches the sample, while theother surface (that does not touch the sample) of the plate is termed“outer surface”.

The term “height” or “thickness” of an object in a QMAX process refersto, unless specifically stated, the dimension of the object that is inthe direction normal to a surface of the plate. For example, spacerheight is the dimension of the spacer in the direction normal to asurface of the plate, and the spacer height and the spacer thicknessmeans the same thing.

The term “area” of an object in a QMAX process refers to, unlessspecifically stated, the area of the object that is parallel to asurface of the plate. For example, spacer area is the area of the spacerthat is parallel to a surface of the plate.

The term of QMAX device refers the device that perform a QMAX (e.g.CROF) process on a sample, and have or not have a hinge that connect thetwo plates.

In some embodiments of QMAX cards, they do not use spacers to controlthe sample thickness in a closed configuration of the movable plates,rather they use other ways to measure the sample thickness afterreaching a closed configuration. The thickness measurements includelight interference measurements.

-   C. Chemicals for Colorimetric Assays

As used herein the term “colorimetric” and grammatical variants thereofrefer to the physical description and quantification of the colorspectrum including the human color perception spectrum (e.g., visiblespectrum). In some embodiments, a colorimetric assay is particularlyuseful when quantification is not necessary and where expensivedetection equipment is unavailable. In certain embodiments, detection ofthe color change can be carried out by naked eye observation of a user(e.g., the person performing the assay). Because a colorimetric assaycan be detected by naked eye observation, a user can either examine thereaction for a detectable change in color or the assay can be carriedout in parallel with one or more controls (positive or negative) thatreplicate the color of a comparable reaction. In some embodiments,calibrated colorimetric measurements could be used to determine theamount of target quantitatively.

In general, a colorimetric analysis involves determining thepresence/absence, level, or concentration of an analyte (such as achemical element or chemical compound) in a sample, such as a solution,with the aid of a color reagent. It is applicable to both organiccompounds and inorganic compounds and may be used with or without anenzymatic reaction step. Generally, the equipment required is acolorimeter, one or more cuvettes, and a suitable color reagent. Theprocess may be automated, e.g., by the use of an AutoAnalyzer or by Flowinjection analysis. In particular embodiments, colorimeters can beadapted for use with plate readers to speed up analysis and reduce thewaste stream.

In one aspect, a colorimetric assay disclosed herein is a non-enzymaticmethod. For example, a metal ion can react with one or more agents toform one or more colored products. For instance, calcium can react witho-cresolphthalein complex one to form a colored complex; copper mayreact with bathocuproin disulfonate to form a colored complex;creatinine can react with picrate to form a colored complex; iron canreact with bathophenanthroline disulfonate to form a colored complex;and phosphate can react with ammonium molybdate and/or ammoniummetavanadate to form a colored complex.

In another aspect, a colorimetric assay disclosed herein comprises oneor more enzymatic reaction step. Typically, the color reaction ispreceded by a reaction catalyzed by an enzyme. As the enzyme is specificto one or more particular substrates, more accurate results can beobtained. For example, in an assay for cholesterol detection such as theCHOD-PAP method), cholesterol in a sample is first reacted with oxygen,catalyzed by the enzyme cholesterol oxidase), to produce cholestenoneand hydrogen peroxide. The hydrogen peroxide is then reacted with4-aminophenazone and phenol, this round catalyzed by a peroxidase, toproduce a colored complex and water. Another example is the GOD-Peridmethod for detecting glucose, where glucose is a sample is first reactedwith oxygen and water, catalyzed by the enzyme glucose oxidase, togenerate gluconate and hydrogen peroxide. The hydrogen peroxide sogenerated then reacts with ABTS to produce a colored complex, and thereaction can be catalyzed by a peroxidase. In yet another example, theso-called GPO-PAP method detects triglycerides, which are firstconverted to glycerol and carboxylic acid (catalyzed by an esterase);the glycerol is then reacted with ATP to form glycerol-3-phosphate andADP (catalyzed by a glycerol kinase); the glycerol-3-phosphate is thenoxidized by a glycerol-3-phosphate oxidase to form dihydroxyacetonephosphate and hydrogen peroxide; and the final enzymatic reaction iscatalyzed by a peroxidase, where the hydrogen peroxide reacts with4-aminophenazone and 4-chlorophenol to form a colored complex. In someembodiments, the colorimetric assay may comprise both non-enzymaticstep(s) and enzymatic step(s). For example, urea can be detected byfirst converting the analyte into ammonium carbonate (catalyzed by aurease), and then the ammonium carbonate reacts with phenol andhypochlorite in a non-enzymatic reaction to form a colored complex.

In some embodiments, a colorimetric assay detects a protein target. Inone aspect, a colorimetric assay involves the formation of aprotein-metal chelation (such as protein-copper chelation), followed bysecondary detection of the reduced metal (e.g., copper). Examples ofthis type of colorimetric assay include the BCA assay and the Lowryprotein assay, such as the Thermo Scientific Pierce BCA and ModifiedLowry Protein Assays. In another aspect, a colorimetric assay involvesprotein-dye binding with direct detection of the color change associatedwith the bound dye. Examples of this type of colorimetric assay includethe 660 nm assay and the Coomassie (Bradford) protein assay. Otherexamples of colorimetric assays for detecting a polypeptide or proteintarget include the Biuret assay, the Bicinchoninic Acid (Smith) assay,the Amido Black method, and the Colloidal Gold assay.

In particular embodiments, the colorimetric assay, such as acolorimetric screening, can be based on NAD(P)H generation. Theabsorbance of NAD(P)H at 340 nm is commonly used to measure the activityof dehydrogenases. Typically, this type of colorimetric assay involvesan indirect method requiring either a synthetic compound or a secondaryenzyme. For example, tetrazolium salts such as nitroblue tetrazolium(NBT) can be reduced to formazan dyes, which absorb light in the visibleregion. These reactions are essentially irreversible under biologicalconditions and the increase in color can be easily monitored visually onfilter discs or on a standard 96-well plate reader. A cascade reactionleading to the formation of a colored formazan links the production ofNAD(P)H to the catalytic activity of a dehydrogenase in a sample.

In particular embodiments, the colorimetric assay is an Enzyme-LinkedImmunosorbent Assay (ELISA). Examples of colorimetric ELISA substratesinclude colorimetric (also called chromogenic) substrate for alkalinephosphatase (AP) and/or horseradish peroxidase enzyme (HRP), such asPNPP (p-Nitrophenyl Phosphate, a widely used substrate for detectingalkaline phosphatase in ELISA applications to produce a yellowwater-soluble reaction product that absorbs light at 405 nm), ABTS(2,2′-Azinobis [3-ethylbenzothiazoline-6-sulfonic acid]-diammonium salt,which is used to detect HRP and yields a water-soluble green endreaction product), OPD (o-phenylenediamine dihydrochloride, which isused to detect HRP and yields a water soluble yellow-orange reactionproduct), and TMB (3,3′,5,5′-tetramethylbenzidine, which yield a bluecolor when detecting HRP).

Specific examples of a colorimetric assay include the HRP/ABTS/H2O2Assay, HRP/4CN/H2O2 Assay, the D-Amino Acid Oxidase Assay, thePeroxidase/o-Dianisidine Assay, the ABTS and o-Dianisidine Assay, theTMB Assay, the Guaiacol Assay, the MNBDH Assay, assays based on theGibbs' Reagent and 4-Aminoantipyrine, the Poly R-478 Assay, theHorseradish Peroxidase-coupled Assay, the MTT assay, the Indole Assay,and the para-Nitrophenoxy Analog (pNA) Assay.

The devices and methods described above may be used to perform any oneor more of the following colorimetric assays. Suitable colorimetricassays include, but are not limited to, colorimetric assays that detectproteins, nucleic acids, antibodies, or microorganisms. Colorimetricassays may be used to determine the concentration of a substance in asolution. In some cases, the colorimetric assays include colorimetricimmunoassays. Suitable colorimetric assays may include those describedin Jiang et al., Analyst (2016), 141: 1196-1208; Morbioli et al., Anal.Chim. Acta. (2017), 970: 1-22; Gu et al., Biotechnology Advances (2015),33: 666-690; Marin et al., Analyst. (2015), 140(1): 59-70; Du et al.,Small. (2013), 9 (9-10): 1467-81; Song et al., Adv. Mater. (2011), 23(37):4215-36; Liu et al., Nanoscale (2011), 3(4):1421-33; Martin et al.J. Animicrob Chemother. (2007) 59 (2): 175-83; Sapan et al. Biotechnol.Appl. Biochem. (1999), 29(pt 2): 99-108.

Colorimetric immunoassays can include enzyme immunoassays such as, e.g.,an enzyme-linked immunosorbent assay (ELISA). ELISA assays can includelabeling a surface bound antigen with an enzyme, e.g., with a singleantibody conjugate or two or more antibodies working in concert to labelthe antigen with the enzyme. An antigen may be immobilized on a solidsurface by non-specific means (e.g., adsorption) or by specific means(e.g., capture by an antibody, in a “sandwich” ELISA). The incubationcan be followed by washing steps and the addition of a detectionantibody covalently linked to an enzyme. In some cases, the detectionantibody is a primary antibody that is itself detected by a secondaryantibody linked to an enzyme. Following labeling of the enzyme, andtypically after one or more washing steps, the enzyme is reacted with anappropriate substrate, such as a chromogenic substrate, in such a manneras to produce a signal, e.g., a chemical signal, that may be detected,e.g., by spectrophotometric, fluorimetric or by visual means. Such colorchange may indicate the presence and/or quantity of the antigen in thesample. Types of ELISA assays include, for example, direct ELISA,sandwich ELISA, and competitive ELISA.

Suitable enzymes for use in enzyme immunoassays include, but are notlimited to, malate dehydrogenase, staphylococcal nuclease,delta-5-steroid isomerase, yeast alcohol dehydrogenase,alpha-glycerophosphate, dehydrogenase, triose phosphate isomerase,horseradish peroxidase, alkaline phosphatase, asparaginase, glucoseoxidase, beta-galactosidase, ribonuclease, urease, catalase,glucose-6-phosphate dehydrogenase, glucoamylase andacetylcholinesterase. The detection in such assays can be accomplishedby colorimetric methods which employ a chromogenic substrate for theenzyme, where suitable substrates include, but are not limited to:o-phenylenediamine (OPD), 3,3′,5,5′-tetramethylbenzidine (TMB),3,3′-diaminobenzide tetrahydrochloride (DAB),2,2′-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid) (ABTS), and thelike. A fluid composition of the substrate, e.g., an aqueous preparationof the substrate, is typically incubated with the substrate surface fora period of time sufficient for the detectable product to be produced.Incubation typically lasts for a period of time ranging from about 10sec to 2 hours, usually from about 30 sec to 1 hour and more usuallyfrom about 5 min to 15 min at a temperature ranging from about 0 to 37°C., usually from about 15 to 30° C. and more usually from about 18 to25° C.

Colorimetric immunoassays can include lateral flow assays (LFA) orimmunochromatography assays. Such assays may be performed on a series ofcapillary beds, e.g., porous paper or polymers, for transporting fluid.Conventional lateral flow test strips include a solid support on which asample receiving area and the target capture zones are supported. Thesolid support material is one which is capable of supporting the samplereceiving area and target capture zones and providing for the capillaryflow of sample out from the sample receiving area to the target capturezones when the lateral flow test strip is exposed to an appropriatesolvent or buffer, which acts as a carrier liquid for the sample.General classes of materials which may be used as supports includeorganic or inorganic polymers, and natural and synthetic polymers. Morespecific examples of suitable solid supports include, withoutlimitation, glass fiber, cellulose, nylon, crosslinked dextran, variouschromatographic papers and nitrocellulose.

At the capture zones, capture molecules may bind the complex, producinga color change in the test strip. The capture zones may include one ormore components of a signal producing system. The signal producingsystem may vary widely depending on the particular nature of the lateralflow assay and may be any directly or indirectly detectable label.Suitable detectable labels for use in the LFA include any moiety that isdetectable by spectroscopic, photochemical, biochemical, immunochemical,electrical, optical, chemical, or other means. For example, suitablelabels include biotin for staining with labeled streptavidin conjugate,fluorescent dyes (e.g., fluorescein, Texas red, rhodamine, greenfluorescent protein, and the like), radiolabels (e.g. ³H, ¹²⁵I, ³⁵S, ¹C,or ₃₂), enzymes (e.g., horseradish peroxidase, alkaline phosphatase andothers commonly used in an ELISA), and colorimetric labels such ascolloidal gold nanoparticles, silver nanoparticles, magneticnanoparticles, cerium oxide nanoparticles, carbon nanotubes, grapheneoxide, conjugated polymers, or colored glass or plastic (e.g.,polystyrene, polypropylene, latex beads). Radiolabels can be detectedusing photographic film or scintillation counters, fluorescent markerscan be detected using a photodetector to detect emitted light. Enzymaticlabels are typically detected by providing the enzyme with a substrateand detecting the reaction product produced by the action of the enzymeon the substrate, and colorimetric labels are detected by simplyvisualizing the colored label.

In some cases, the colorimetric assay may be used to measure ions in asample. For example, chloride ions can be measured by a colorimetricassay. Chloride ions displace thiocyanate from mercuric thiocyanate.Free thiocyanate reacts with ferric ions to form a coloredcomplex—ferric thiocyanate, which is measured photometrically.

Likewise, magnesium can be measured colorimetrically using calmagite,which turns a red-violet color upon reaction with magnesium; by aformazan dye test; emits at 600 nm upon reaction with magnesium or usingmethylthymol blue, which binds with magnesium to form a blue coloredcomplex.

Likewise, calcium can be detected by a colorimetric technique usingO-Cresolphtalein, which turns a violet color upon reaction of0-Cresolphtalein complexone with calcium.

Likewise, bicarbonate can be tested bichromatically because bicarbonate(HCO3⁻) and phosphoenolpyruvate (PEP) are converted to oxaloacetate andphosphate in the reaction catalyzed by phosphoenolpyruvate carboxylase(PEPC). Malate dehydrogenase (MD) catalyzes the reduction ofoxaloacetate to malate with the concomitant oxidation of reducednicotinamide adenine dinucleotide (NADH). This oxidation of NADH resultsin a decrease in absorbance of the reaction mixture measuredbichromatically at 380/410 nm proportional to the Bicarbonate content ofthe sample. Blood urea nitrogen can be detected in a colorimetric testin which diacetyl, or fearon develops a yellow chromogen with urea andcan be quantified by photometry. Likewise, creatinine can be measuredcolorimetrically, by treated the sample with alkaline picrate solutionto yield a red complex. In addition, creatine can be measured using anon-Jaffe reaction that measures ammonia generated when creatinine ishydrolyzed by creatinine iminohydrolase. Glucose can be measured in anassay in which blood is exposed to a fixed quantity of glucose oxidasefor a finite period of time to estimate concentration. After thespecified time, excess blood is removed and the color is allowed todevelop, which is used to estimate glucose concentration. For example,glucose oxidase reaction with glucose forms nascent oxygen, whichconverts potassium iodide (in the filter paper) to iodine, forming abrown color. The concentration of glycosylated hemoglobin as an indirectread of the level of glucose in the blood.

Plasma high-density lipoprotein cholesterol (HDL-C) determination ismeasured by the same procedures used for plasma total cholesterol, afterprecipitation of apoprotein B-containing lipoproteins in whole plasma(LDL and VLDL) by heparin-manganese chloride. These compounds can alsobe detected colorimetrically in an assay that is based on the enzymedriven reaction that quantifies both cholesterol esters and freecholesterol. Cholesterol esters are hydrolyzed via cholesterol esteraseinto cholesterol, which is then oxidized by cholesterol oxidase into theketone cholest-4-en-3-one plus hydrogen peroxide. The hydrogen peroxideis then detected with a highly specific colorimetric probe. Horseradishperoxidase catalyzes the reaction between the probe and hydrogenperoxide, which bind in a 1:1 ratio. Samples may be compared to a knownconcentration of cholesterol standard.

-   Examples of Reagents-   A. Glucose Colorimetric (Fluorimetric) Assay-   Sample: Whole Blood, Plasma, Serum, Saliva-   Reagent Recipe 1:-   100 units/ml Glucose Oxidase, 100 unit/ml Horseradish Peroxidase, 20    mM 4-amino antipyrine, 20 mM TOOS-   Reagent Recipe 2:-   100 unit/ml Glucose Oxidase, 100 unit/ml Horseradish Peroxidase, 20    mM 3,5,3′,5′-Tetramethylbenzidine (TMB)-   Reagent Recipe 3:-   100 unit/ml Glucose Oxidase, 100 unit/ml Horseradish Peroxidase, 20    mM Amplex Red-   Reagent Recipe 4:-   1 unit/ml Hexokinase, 220 mg/ml ATP, 400 mg/ml NAD-   B. Calcium Colorimetric Assay-   Sample: Whole Blood, Plasma, Serum, Saliva-   Reagent Recipe 1:-   17 mg/ml Arsenazo III-   C. Albumin Colorimetric Assay-   Sample: Whole Blood, Plasma, Serum, Saliva-   Reagent Recipe 1:-   22 mg/ml Bromcresol purple-   D. Total Protein Colorimetric assay-   Sample: Whole Blood, Plasma, Serum, Saliva-   Reagent Recipe 1:-   1.34 mg/ml Cupric sulfate, 3.43 mg/ml Sodium potassium tartrate,    0.28 mg/ml Potassium iodide-   E. Sodium Colorimetric Assay-   Sample: Whole Blood, Plasma, Serum, Saliva-   Reagent Recipe 1:-   220 mg/ml ONPG, 0.05 unit/ml β-Galactosidase-   F. Potassium Colorimetric Assay-   Sample: Whole Blood, Plasma, Serum, Saliva-   Reagent Recipe 1:-   220 mg/ml ADP, 0.05 unit/ml Phosphoenolpyruvate, 0.1 unit/ml    Pyruvate kinase, 480 mg/ml NADH, 13.6 mg/ml Potassium phosphate, 95    mg/ml Magnesium sulfate, 7.85 mg/ml FAD, 130 mg/ml    4-Aminoantipyrine, 10 unit/ml Horseradish Peroxidase, 1.88 mg/ml    TBHBA-   G. Chloride Colorimetric Assay-   Sample: Whole Blood, Plasma, Serum, Saliva-   Reagent Recipe 1:-   530 mg/ml CNPG3, 0.36 unit/ml α-Amylase, 250 mg/ml Calcium acetate-   H. Blood Urea Nitrogen Colorimetric Assay-   Sample: Whole Blood, Plasma, Serum, Saliva-   Reagent Recipe 1:-   0.5 U/ml Urea Amidolyase, 570 ug/ml PEP, 220 ug/ml ATP, 1 U/ml    Pyruvate Kinase 10U/ml Pyruvate Oxidase, 13.6 mg/ml Potassium    phosphate, 95 ug/ml MgCl2, 7.85 ug/ml FAD-   1.88 mg/ml TBHBA, 130 ug/ml 4-AAP, 10 U/ml Peroxidase-   I. Creatinine Colorimetric Assay-   Sample: Whole Blood, Plasma, Serum, Saliva-   Reagent Recipe 1:-   10 U/ml Creatinine Amidohydrolase, 30 U/ml Creatinine    Amidinohydrolase, 10 U/ml Sarcoosine Oxidase, 1.88 mg/ml TBHBA, 130    ug/ml 4-AAP, 10 U/ml Peroxidase-   J. Alkaline Phosphatase Colorimetric Assay-   Sample: Whole Blood, Plasma, Serum, Saliva-   Reagent Recipe 1:-   560 ug/ml p-Nitrophenyl Phosphate, 0.5 U/ml Zinc Sulfate, 330 ug/ml    Magnesium Sulfate-   K. Alanine Amino Transferase Colorimetric Assay-   Sample: Whole Blood, Plasma, Serum, Saliva-   Reagent Recipe 1:-   8.74 mg/ml L-Alanine, 1.01 mg/ml α-Ketoglutaric Acid-   10U/ml Pyruvate Oxidase, 13.6 mg/ml Potassium phosphate, 95 ug/ml    MgCl2, 7.85 ug/ml FAD, 1.88 mg/ml TBHBA, 130 ug/ml 4-AAP, 10 U/ml    Peroxidase-   L. Aspartate Amino Transferase Colorimetric Assay-   Sample: Whole Blood, Plasma, Serum, Saliva-   Reagent Recipe 1:-   4.26 mg/ml L-Aspartic Acid, 1.01 mg/ml α-Ketoglutaric Acid, 10 U/ml    Oxaloacetate decarboxylase, 1.88 mg/ml TBHBA, 130 ug/ml 4-AAP, 10    U/ml Peroxidase-   M. Bilirubin Colorimetric Assay-   Sample: Whole Blood, Plasma, Serum, Saliva-   Reagent Recipe 1:-   1 U/ml Bilirubin Oxidase-   N. Cholesterol Colorimetric (Fluorimetric) Assay-   Sample: Whole Blood, Plasma, Serum, Saliva-   Reagent Recipe 1:-   100 unit/ml Cholesterol Oxidase, 100 unit/ml Horseradish Peroxidase,    20 mM 4-amino antipyrine, 20 mM TOOS-   Reagent Recipe 2:-   100 unit/ml Cholesterol Oxidase, 100 unit/ml Horseradish Peroxidase,    20 mM 3,5,3′,5′-Tetramethylbenzidine (TMB)-   Reagent Recipe 3:-   100 unit/ml Cholesterol Oxidase, 100 unit/ml Horseradish Peroxidase,    20 mM Amplex Red-   O. Triglycerides Colorimetric (Fluorimetric) Assay-   Sample: Whole Blood, Plasma, Serum, Saliva-   Reagent Recipe 1:-   100 unit/ml Lipase, 100 unit/ml Glycerokinase, 100 unit/ml    Glycerophosphate Oxidase, 20 mM 4-amino antipyrine, 20 mM TOOS-   Reagent Recipe 2:-   100 unit/ml Lipase, 100 unit/ml Glycerokinase, 100 unit/ml    Glycerophosphate Oxidase, 20 mM 3,5,3′,5′-Tetramethylbenzidine (TMB)-   Reagent Recipe 3:-   100 unit/ml Lipase, 100 unit/ml Glycerokinase, 100 unit/ml    Glycerophosphate Oxidase, 20 mM Amplex Red-   P. Alcohol Colorimetric (Fluorimetric) Assay-   Sample: Whole Blood, Plasma, Serum, Saliva, Breath-   Reagent Recipe 1:-   100 unit/ml Alcohol Oxidase, 100 unit/ml Horseradish Peroxidase, 20    mM 4-amino antipyrine, 20 mM TOOS-   Reagent Recipe 2:-   100 unit/ml Alcohol Oxidase, 100 unit/ml Horseradish Peroxidase, 20    mM 3,5,3′,5′-Tetramethylbenzidine (TMB)-   Reagent Recipe 3:-   100 unit/ml Alcohol Oxidase, 100 unit/ml Horseradish Peroxidase, 20    mM Amplex Red-   Q. Hydrogen Peroxide (Fluorimetric) Assay-   Sample: Whole Blood, Plasma, Serum, Saliva, Breath-   Reagent Recipe 1:-   100 unit/ml Horseradish Peroxidase, 20 mM 4-amino antipyrine, 20 mM    TOOS-   Reagent Recipe 2:-   100 unit/ml Horseradish Peroxidase, 20 mM    3,5,3′,5′-Tetramethylbenzidine (TMB)-   Reagent Recipe 3:-   100 unit/ml Horseradish Peroxidase, 20 mM Amplex Red-   R. Gram Staining-   Sample: Blood smear, Vaginal samples, Genital samples-   Gram Crystal Violet-   20 g Crystal Violet, 8 g Ammonium Oxalate, 200 mL Methanol-   Gram Iodine-   3.33 g Iodine Crystal, 6.67 g Potassium Iodide-   Gram Decolorizer-   500.0 mL Ethanol, 500.0 mL Acetone-   Gram Safranin-   0.25 g Safranin O, 10 mL Ethanol-   Gram Basic Fuchsin Basic-   0.7 g Fuchsin, 3.5 mL Phenol, 14 mM Ethanol-   S. Leishman Staining-   Sample: Smear sample-   Recipe 1-   0.2 g Leishman's dye, 100 mL Acetone-free methyl alcohol-   T. Giemsa Staining-   Sample: Smear sample-   Recipe 1-   0.15 g Giemsa powder, 12.5 mL Glycerin, 12.5 mL Methyl alcohol-   U. Wright Staining-   Sample: Smear sample-   Recipe 1-   1.5 g Wright stain, 500 mL Methanol-   V. Field Staining-   Sample: Smear sample-   Field Solution A-   1.6 g Methylene Blue, 10 g Disodium dihydrogen phosphate, 12.5 g    Potassium dihydrogen phosphate, lg Azur, 1000 mL Distilled water-   Field Solution B-   2 g Eosin Y, 10 g Disodium dihydrogen phosphate, 12.5 g Potassium    dihydrogen phosphate,-   1000 mL Distilled water-   W. Jenner Staining-   Sample: Smear sample-   Recipe 1-   0.5 g Jenner stain, 100 mL Methanol-   X. JSB Staining-   Sample: Smear sample-   Recipe 1-   0.5 g Atine orange dye, 3 mL 1% Sulfuric acid, 0.5 g Potassium    dichromate, 3.5 g Disodium hydrogen phosphate dehydrate, 500 mL    Distilled water-   JSB stain II-   1 g Eosin Y, 500 ml Distilled water-   Y. White Blood cells staining for counting and differentiate-   Sample: Blood, urine, other body fluidics-   Recipe 1-   1 ug/mL to 1 mg/mL Acridine Orange (Detection agents)-   Recipe 2-   150 uM Propidium Iodide (PI) (Detection agents), 100 uM Fluorescein    Isothiocyanate (FITC), 250 uM Basic Orange 21 (B021) dye-   Z. Platelets Staining for Counting-   Sample: Blood, urine, other body fluidics-   Recipe 1-   1 ug/mL to 1 mg/mL Acridine Orange (Detection agents)-   Recipe 2-   150 uM Propidium Iodide (PI) (Detection agents), 100 uM Fluorescein    Isothiocyanate (FITC),-   250 uM Basic Orange 21 (B021) dye

Testing System with QMAX Device

One aspect of the present invention provides systems and methods ofanalyzing a bio/chemical sample using QMAX device.

-   AA1. A method for analyzing a sample, comprising:    -   a) depositing a sample on a Q-card and closing the Q-card;    -   b) inserting the closed Q-card into an adaptor that connects to        a camera of a handheld mobile communication device;    -   c) taking image(s) of the closed Q-card using the camera of the        handheld mobile communication device;    -   d) transmitting, to a remote location, the image(s) and/or an        analysis result of the images from the handheld mobile        communication device;    -   e) analyzing, at the remote location, the image(s) and/or the        analysis result transmitted from the mobile communication        device; and    -   f) notifying a third party and/or the handheld mobile        communication device if an anomaly is detected;

wherein the Q-card comprises two plates that are movable relative toeach other and have an open configuration and a closed configuration;

wherein the sample is deposited on one or both plates of the Q-Card atthe open configuration, and at the closed configuration at least a partof the sample is between the two plates,

wherein the mobile communication device is configured to produce animage of the Q card in the adaptor and transmit the image and/or ananalysis result of the same to a remote location.

-   AA2. The method of any prior embodiment, wherein the sample    deposited onto the Q-card is from a subject, and the subject    performs step a).-   AA3. The method of any prior embodiment, wherein the anomaly is    identified if the analysis result of the sample is not within a    normal range.-   AA4. The method of any prior embodiment, wherein the anomaly is    identified if the analysis results produced by the remote device and    the mobile handheld communication device differ by a pre-defined    value.-   AA5. The method of any prior embodiment, wherein the sample    comprises a body fluid selected from the group consisting of:    amniotic fluid, aqueous humour, vitreous humour, blood (e.g., whole    blood, fractionated blood, plasma, serum, etc.), breast milk,    cerebrospinal fluid (CSF), cerumen (earwax), chyle, chime,    endolymph, perilymph, feces, gastric acid, gastric juice, lymph,    mucus (including nasal drainage and phlegm), pericardial fluid,    peritoneal fluid, pleural fluid, pus, rheum, saliva, sebum (skin    oil), semen, sputum, sweat, synovial fluid, tears, vomit, urine and    exhaled condensate.-   AA6. The method of any prior embodiment, wherein the sample    comprises an environmental specimen that is obtained from: river,    lake, pond, ocean, glaciers, icebergs, rain, snow, sewage,    reservoirs, tap water, drinking water, soil, compost, sand, rocks,    concrete, wood, brick, sewage; air, heat vents, industrial exhaust,    or vehicular exhaust.-   AA7. The method of any prior embodiment, wherein the sample    comprises a foodstuff specimen that includes: raw food ingredients,    cooked or processed food, plant and animal sources of food,    preprocessed food, or fully processed food.-   AA8. The method of any prior embodiment, wherein, in step (a), the    Q-card is pressed by human hand.-   AA9. The method of any prior embodiment, wherein step e) comprises    comparing the result to a threshold or normal range to identify    samples that contain an anomaly.-   AA10. The method of any prior embodiment, wherein the method further    comprises updating the handheld mobile communication device if the    analysis at the remote location produces a result that is    significantly different.-   AA11. The method of any prior embodiment, wherein the sample    deposited onto the Q-card is from a subject, and the analysis result    is not transmitted to the subject.-   AA12. The method of any prior embodiment, wherein the third party is    a medical professional.-   AA13. The method of embodiment AA12, wherein the medical    professional is a doctor or nurse practitioner.-   AA14. The method of any of embodiments AA1-AA12, wherein third party    is an insurance company.-   AA15. The method of any prior embodiment, wherein the result from    the mobile communication device and/or the result from the remote    location are sent to an emergency room.-   AA16. The method of embodiment AA1, wherein, based on the results,    the handheld mobile communication device or the remote location    transmits follow-up information to the subject.-   AA17. The method of embodiment AA16, wherein the follow-up    information comprises an explanation of the result, education about    a disease or condition, information related to a possible treatment,    information on the location of a suitable physician, information    related to change of diet and/or exercises, or an advertisement.-   AA18. The method of any prior embodiment, wherein the Q-card    comprises spacers that have a substantially uniform height and a    predetermined constant inter spacer distance, and in the closed    configuration: at least part of the sample is compressed by the two    plates of the Q-card into a layer of highly uniform thickness and is    substantially stagnant relative to the plates, wherein the uniform    thickness of the layer is confined by the inner surfaces of the two    plates and is regulated by the plates and the spacers.-   AA19. The method of embodiment AA18, wherein at least one of the    plates is flexible.-   AA20. The method of embodiment AA19, wherein for the flexible plate,    the thickness of the flexible plate times the Young's modulus of the    flexible plate is in the range 60 to 750 GPa-um.-   AA21. The method of embodiment AA19, wherein for the flexible plate,    the fourth power of the inter-spacer-distance (ISD) divided by the    thickness of the flexible plate (h) and the Young's modulus (E) of    the flexible plate, ISD4/(hE), is equal to or less than 106 um3/GPa,-   AA22. The method of embodiment AA18, wherein spacers regulating the    layer of uniform thickness have a filling factor of at least 1%,    wherein the filling factor is the ratio of the spacer area in    contact with the layer of uniform thickness to the total plate area    in contact with the layer of uniform thickness.-   AA23. The method of embodiment AA18, wherein for spacers regulating    the layer of uniform thickness, the Young's modulus of the spacers    times the filling factor of the spacers is equal or larger than 10    MPa, wherein the filling factor is the ratio of the spacer area in    contact with the layer of uniform thickness to the total plate area    in contact with the layer of uniform thickness.-   AA24. The method of any prior embodiment, wherein one or both plates    comprises a location marker, either on a surface of or inside the    plate, that provide information of a location of the plate.-   AA25. The method of any prior embodiment, wherein one or both plates    comprises a scale marker, either on a surface of or inside the    plate, that provide information of a lateral dimension of a    structure of the sample and/or the plate.-   AA26. The method of any prior embodiment, wherein one or both plates    comprises an imaging marker, either on surface of or inside the    plate, that assists an imaging of the sample.-   AA27. The method of embodiment AA18, wherein the spacers functions    as a location marker, a scale marker, an imaging marker, or any    combination of thereof.-   AA28. The method of embodiment AA18, wherein the average thickness    of the layer of uniform thickness is in the range of 0.2 μm to 3.8    μm and the sample is blood.-   AA29. The method of embodiment AA18, wherein the inter-spacer    distance is in the range of 7 μm to 50 μm.-   AA30. The method of embodiment AA18, wherein the inter-spacer    distance is in the range of 50 μm to 120 μm.-   AA31. The method of embodiment AA18, wherein the inter-spacer    distance is in the range of 120 μm to 200 μm.-   AA32. The method of embodiment AA18, wherein the inter-spacer    distance is substantially periodic.-   AA33. The method of embodiment AA18, wherein the spacers are pillars    with a cross sectional shape selected from round, polygonal,    circular, square, rectangular, oval, elliptical, or any combination    of the same.-   AA34. The method of embodiment AA18, wherein the spacers have are    pillar shape and have a substantially flat top surface, wherein, for    each spacer, the ratio of the lateral dimension of the spacer to its    height is at least 1.-   AA35. The method of embodiment AA18, wherein each spacer has the    ratio of the lateral dimension of the spacer to its height is at    least 1.-   AA36. The method of embodiment AA18, wherein the minimum lateral    dimension of spacer is less than or substantially equal to the    minimum dimension of an analyte in the sample.-   AA37. The method of embodiment AA18, wherein the minimum lateral    dimension of spacer is in the range of 0.5 um to 100 um.-   AA38. The method of embodiment AA18, wherein the spacers have a    pillar shape, and the sidewall corners of the spacers have a round    shape with a radius of curvature at least 1 μm.-   AA39. The method of embodiment AA18, wherein the spacers have a    density of at least 1000/mm2.-   AA40. The method of any prior embodiment, wherein at least one of    the plates is transparent.-   AA41. The method of any prior embodiment, wherein at least one of    the plates is made from a flexible polymer.-   AA42. The method of embodiment AA18, wherein, for a pressure that    compresses the plates, the spacers are not compressible and/or,    independently, only one of the plates is flexible.-   AA43. The method of any prior embodiment, wherein the flexible plate    has a thickness in the range of 10 um to 200 um.-   AA44. The method of embodiment AA18, wherein the variation of the    uniform thickness is less than 30%.-   AA45. The method of embodiment AA18, wherein the variation of the    uniform thickness is less than 10%.-   AA46. The method of embodiment AA18, wherein the variation of the    uniform thickness is less than 5%.-   AA47. The method of any prior embodiment, wherein the plates are    connected by a hinge and are configured to be changed from the open    configuration to the closed configuration by folding the plates    along the hinge.-   AA48. The method of any prior embodiment, wherein the layer of    uniform thickness sample is uniform over a lateral area that is at    least 1 mm2.-   AB 1. A system for analyzing a sample, comprising:    -   a) a Q-card for manipulating a sample for analysis comprising        two plates that are movable relative to each other and have an        open configuration and a closed configuration;    -   b) a handheld mobile communication device that comprises a        camera;    -   c) an adaptor having a slot that is configured to hold a closed        Q-Card, wherein the adaptor connects to the handheld mobile        communication device and permits the camera to take an image of        closed Q-Card; and    -   d) a remote device that is capable of storing information and        communicating with the mobile communication device;        -   wherein the sample is deposited on one or both plates of the            Q-Card at the open configuration, and at the closed            configuration at least a part of the sample is between the            two plates,        -   wherein the system is configured to produce an image of the            Q card in the adaptor and transmit the image and/or an            analysis result of the same to a remote location.-   AB2. The system of embodiment AB1, wherein the Q-card can be placed    in the closed configuration by folding.-   AB3. The system of embodiment AB1, wherein the remote device is    configured to analyze the image and/or the analysis result of the    same.-   AB4. The system of embodiment AB1, wherein the remote device is    configured to communicate with other remote devices.-   AB5. The system of embodiment AB1, wherein the remote device is    configured to notify a third if an anomaly in a sample placed in the    Q card is detected.-   AC1. A method for providing healthcare recommendations to a subject,    comprising:    -   a) using Q-cards and an associated mobile communication device        to analyze one or a plurality of analytes in samples from a        subject;    -   b) transmitting, to a remote location, the analysis results of        the analytes from the mobile communication device;    -   c) storing the analysis results in a data set;    -   d) generating, at the remote location, a series of healthcare        recommendations based on accumulated analysis results in the        data set; and    -   e) providing the healthcare recommendations to the subject by        sending messages to the mobile communication device;

wherein the healthcare recommendations comprise suggestions related tomedicine, nutrition/diet, exercise, and/or treatment for the subject.

-   AC2. The method of paragraph AC1, further comprising identifying the    subject's needs before providing the healthcare recommendations to    the subject.

B. Cholesterol Testing with QMAX Device

Another aspect of the present invention provides devices and methods ofcholesterol testing using QMAX device.

-   BA1. A method of analyzing a liquid sample, comprising:    -   (a) obtaining the liquid sample;    -   (b) obtaining a device, which comprises a first plate, a second        plate, and spacers fixed on one or both of the plates; wherein:        -   i. the plates are movable relative to each other into            different configurations, including an open configuration            and a closed configuration;        -   ii. each plate respectively comprises an inner surface that            has a sample contact area, and        -   iii. the spacers have a predetermined substantially uniform            height, and at least one of the spacers is inside the sample            contact area;    -   (c) depositing the sample on one or both of the plates when the        plates are in an open configuration,        -   wherein in the open configuration the two plates are            partially or entirely separated apart and the spacing            between the plates is not regulated by the spacers; and    -   (d) after (c), bringing the two plates together and pressing the        plates into a closed configuration,        -   wherein in the closed configuration: at least part of the            sample is compressed by the two plates into a layer of            highly uniform thickness, which is confined by the inner            surfaces of the two plates and is regulated by the spacers;        -   wherein one or both sample contact surfaces comprise one or            more storage sites that store one or more reagents, which            are configured to dissolve and diffuse in the sample in the            closed configuration, and react with cholesterol in the            sample to produce or alter a luminescence signal;    -   (e) reading the luminescence signal from the layer of highly        uniform thickness, thereby obtaining a measurement of total        cholesterol in the sample.-   BA2. The method of paragraph BA1, wherein the one or more reagents    are configured to react with cholesterol to generate or alter a    colormetric luminescence signal, wherein the reading step (e)    comprises detecting and quantifying the colormetric luminescence    signal from the analyte in the layer of highly uniform thickness.-   BA3. The method of paragraph BA1, wherein the one or more reagents    comprise cholesteryl ester hydrolase and cholesterol oxidase.-   BA4. The method of paragraph BA3, wherein the one or more reagents    further comprise peroxidase and a color probe.-   BA5. The method of paragraph BA4, wherein the color probe comprises    4-aminophenazone and phenol.-   BA6. The method of paragraph BA1, wherein the one or more storage    sites comprise a first storage site located on the first plate and a    second storage site located on the second plate.-   BA7. The method of paragraph BA6, wherein:    -   i. the first storage site comprises cholesteryl ester hydrolase        and cholesterol oxidase; and    -   ii. the second storage site comprises 4-aminophenazone, phenol        and peroxidase.

C. Heavy Metal Testing

Another aspect of the present invention provides devices and methods ofheavy metal testing in bio/chemical samples. More specifically, theinvention provides a process for detecting heavy metal ions in anaqueous system, a device comprising the heavy metal ion test piece and asensor. A portable test method provided by the device according to theinvention, so as to detect the heavy metal ions in a convenient,efficient and rapid manner.

The heavy metal (ion) pollution refers to the environmental pollutioncaused by heavy metals or their compounds. The increase of the heavymetal content in the environment, especially in the case of heavy metalpollution in an aqueous system, is mainly due to human factors, such asmining, waste gas emission, sewage irrigation and the use of heavymetal-containing products, which results in the deterioration ofenvironmental quality. Currently there is still a need for a heavy metalion test piece which can be used to detect the small amount, even traceamount of heavy metal ions in an aqueous system in a simple, low cost,highly sensitive, highly reliable and stable manner. Meanwhile, it isrequired that the test piece is available for in situ detection, and iscapable of detecting heavy metal ions with high sensitivity. Moreover,it is desired that the heavy metal ions can be not only qualitativelydetected, but also quantitatively or semi-quantitatively detected. Thecurrent invention provides devices and methods for achieving thesegoals.

C-1. Devices and Methods for Heavy Metal Testing

FIG. C1 shows that the invention comprises two parts: 1. Test, whichcomprises a test card that has dried reagent in a volume-controlledsample chamber, and can be inserted into a smartphone-based reader formeasurement; 2. Calculation, which comprises a method to convert thephotograph taken by smartphone and convert to signal for calculatinganalyte concentrations.

As demonstrated by FIG. C1 , this invention is a device and method forobtaining a point-of-collection, selected quantitative indicia of ananalyte on a test platform, comprising:

-   -   1. providing a modular, colorimetric reactive test platform        having a test region and calibration region;    -   2. providing an analyte to be tested on the test region of the        modular, colorimetric test platform, wherein the test region is        adapted to enable a colorimetric reaction to the analyte;    -   3. obtaining a color image of the test region containing the        analyte and the calibration region;    -   4. selecting an array of pixels in each of the color images of        the test region containing the analyte and the calibration        region;    -   5. determining a median RGB color value for each of the arrays        of pixels;    -   6. converting the median RGB color value for each of the arrays        of pixels to a characteristic value;    -   7. providing a calibration indicia that relates a selected        quantitative indicia of the characteristic value;    -   8. associating the characteristic value to determine the        selected quantitative indicia of the analyte

As shown in FIG. C2 , a first plate, which is a coerce white substrate,is printed uniformly with color indicator as well as pH regulatingagent. The color indicator is bio/chemical reagent that shows specificreaction to heavy metals in liquid sample. The liquid sample includes,but is not limited to, water, soil sample, oil, body fluid and food. Incertain embodiments, the sample is drinking water. In certainembodiments, the sample is food. In some embodiments, the first plate isa coerce white polystyrene plate. In some embodiments, the colorindicator is dried on the first plate. In some embodiments, the pHregulating agent is dried on the first plate. In some embodiments, theconcentration of dried color indicator is 1 uM to 10 mM. In someembodiments, the concentration of dried pH regulating agent is 1 uM to10 mM.

As shown in FIG. C2 , the surface of the first plate facing the secondplate is defined as the inner surface of the first plate; the surface ofthe second plate that faces the first plate are also defined as theinner surface of the second plate. In some embodiments, the innersurfaces of the respective plates comprise a sample contact area forcontacting a sample that comprises an analyte. The sample contact areacan occupy part or the entirety of the respective inner surface.

As shown in FIG. C2 , for testing heavy metal in water usingcolorimetric tests, a pH regulating agent must add to the sample toadjust the pH level to optimum condition. This is because the chemicalreaction rate of color indicator to heavy metal ions changessignificantly at different pH level, which leads to large colorvariation within tests if the pH is unregulated. For heavy metal test, apH regulating agent, or a combination of multiple combination of them,is dried on the plate for adjusting sample PH level includes, but is notlimited to: Formic acid (methanoic acid), Oxalic acid (ethanedioicacid), Lactic acid (2-hydroxypropanoic acid), Malic acid(2-hydroxybutanedioic acid), Citric acid(2-hydroxypropane-1,2,3-tricarboxylic acid), Carbonic acid(hydroxymethanoic acid, not an IUPAC name), Aminomethylphosphonic acid.

As shown in FIG. C2 , the second plate comprises spacers that are fixedon the inner surface of the second plate. It should be noted, however,that in some embodiments the spacers are fixed on the inner surface ofthe first plate and in other embodiments on the inner surfaces of boththe second plate and the first plate

As shown in FIG. C2 , the spacer is between 1 um, 2 um, 5 um, 10 um, 20um, 50 um, 100 um, 200 um, 500 um, 1000 um or in a range between any ofthe two values. The diameter of hole in the spacer is around 0.5 mm, 1mm, 2 mm, 3 mm 4 mm, 5 mm, or in a range between any of the two values.The center-to-center spacing between holes is 1 mm, 2 mm, 3 mm, 4 mm, 5mm, 6 mm, 7 mm, 8 mm, 9 mm, 10 mm, 20 mm, 50 mm. or in a range betweenany of the two values. The second plate is a transparent flat film, withthickness around 1 um, 2 um, 5 um, 10 um, 20 um, 50 um, 100 um, 200 um,500 um, 1000 um or in a range between any of the two values.

As shown in FIG. C2 , the first plate and the second plate are moveablerelative to each other into different configuration. One of theconfigurations is an open configuration, in which the two plates arepartially or entirely separated apart and the spacing between the platesare not regulated by the spacers. FIG. C1 shows the plates in the openconfiguration, in which a sample, such as but not limited to blood, canbe added to first plate, the second plate, or both of the plates. Insome embodiments, the inner surface of a respective plate comprises asample contact area, which occupies a part of the entirety of the innersurface. In certain embodiments, the spacers are positioned within thesample contact area. In some embodiments, the spacers are not fixed toany one of the plates, but are mixed in the sample.

As shown in FIG. C2 The second plate is a transparent thin film withsmooth surface. It is necessary that the absorption of second plate doesnot interfere with the absorption of color indicator. Depends on theflexibility of the material, thickness from 10 um˜300 um can be used assecond plate, as long as no distortion of sample chamber will happenafter second plate is pressed onto the sample.

FIG. 3 shows a Schematics of test procedure. 1. First, minute samplesare added to each well printed with color indicator and and pHregulating agent. 2. The transparent second plate is then pressed on topof the spacer to form a closed sample chamber. 3. Incubation about 1 minto allow each individual sample to develop color. In this process, thecolor indicator and pH regulating agent is fully dissolved and mixed.

As shown in FIG. C3 , a white polystyrene (PS) substrate printed withhome-made color indicator and pH regulating agent. The color indicatorand pH regulating agent amount on the sensing area is carefullycontrolled according to the dimension of the well, so that when eachwell is filled full with sample, the desired pH level and colorindicator concentration can be achieved. Depends on the type of heavymetal or their combinations, different chemicals are used as colorindicator. Color Indicator can be: (1) For lead detection, the colorindicator is 0.01%˜0.2% Sodium Rhodizonate (preferable 0.2% afterdissolved in sample), or (2) For Copper, Cadmium, Chromium, Mercury, 10uM˜1 mM Dithizone (preferable 30 uM after dissolved in sample)

As shown in FIG. C3 , the printing parameter for Color Indicator agentcan vary as long as uniform drying is achieved on the first plate. Theprinting conditions, i.e., droplet volume, speed, depends on the surfacewetting property of the first plate, which is well-known to skilledperson, thus do not require elucidation. In this invention, the printingcondition is droplet diameter 500˜600 um, pitch˜1 mm, print speed˜10mm/sec.

As shown in FIG. C3 , the well dimension is determined by dimensions ofholes array on the spacer. The thickness of the spacer, the diameter ofthe holes and their spacing determines the sample volume. Theirconfiguration is flexible but it is crucial to avoid distortion ofsample chamber under certain configurations, i.e. small aspect ratio.Here, the thickness of the spacer can be 2 um˜1 mm (preferably 100 um),and the well diameter can be 100 um˜10 mm (preferably, 3 mm), and thecenter-to-center spacing can be 100 um˜10 mm, (preferably, 6 mm).

As shown in FIG. C3 , In some embodiments, the method of the presentinvention, after step (2) and before step (3), further compriseincubating the layer of uniform thickness for a predetermined period oftime. In certain embodiments, the predetermined period of time is equalto or longer than the time needed for the detection antibody to diffuseinto the sample across the layer of uniform thickness. In certainembodiments, the predetermined period of time is less than 10 seconds,20 seconds, 30 seconds, 45 seconds, 1 minute, 1.5 minutes, 2 minutes, 3minutes, 4 minutes, 5 minutes, 6 minutes, 7 minutes, 8 minutes, 9minutes, 10 minutes, 15 minutes, 20 minutes, 30 minutes, or 60 minutes,or in a range between any of the two values.

FIG. C4 shows the diagram of a chemical reaction that is used to testlead in water. The lead ion reacts with Sodium Rhodizonate (dark yellowcolor) dissolved in sample, which form a insoluble lead Rhodizonate thathas a red-crimson color. The color absorption can be analyzed tocalculate the lead concentration in water.

FIG. C5 A diagram of a chemical reaction that is used to test heavymetals in water. The heavy metals can be Cd, Cu, Cr, Hg. The heavy metalion reacts with Dithiozone dissolved in sample, which form aDithizone-Metal complex that yield a different color for different heavymetals. The color can be used to identify the type of heavy metals andthe color absorption can be analyzed to calculate the heavy metalconcentration in water.

FIG. C6 shows schematics of converting colorimetric Lead in water teststandard curve of individual R, G, B channel to a single standard curve.For each sample contains different concentration of heavy metals, the R,G, B signal are different. A combination of R, G, B channel signal atdifferent Lead concentration is used for this conversion. In someembodiment, the method of combination is linear combination. In someembodiment, the coefficient for combining RGB channel signal, is aconstant. In some embodiment, the coefficient for combining RGB channelsignal, is a matrix. In some embodiment, the coefficient for combiningRGB channel signal, is a function of lead concentration in water.

As shown in FIG. C7 the algorithm to converting standard curve ofindividual R, G, B channel to a single standard curve is a process tofind the best coefficient of combing R,G,B signals so that bestsensitivity of assay can be achieved. In some embodiment, a linearcombination of R, G, B channel signal at different Lead concentration isused for this conversion. In some embodiment, the linear coefficient istrained using a Generalized Reduced Gradient Algorithm. Such algorithmis open source and known to skilled person and does not requireelucidation. Here, the process of this algorithm is shown in a diagram,briefly:

-   -   1. First, we define 4 constant: C1, C2, C3, and C4 so that        Signal=C₁*R+C₂G+C₃*B+C₄    -   2. Change the linear coefficient by a small amount with        pre-defined amount    -   3. Calculate the limit of detection (LOD),    -   4. keep changing the linear coefficient until the minimum LOD        can be achieved

In this invention, we trained the data using 48 different tests. It isexpected that the precision can be further improved with more trainingdata. This well known among skilled person and does not require furtherelucidation.

-   C-2. Example: Test Lead Concentration in Tap Water

As an example, we Prepare a chip for testing lead in water. On a whitecoerce PS substrate we printed with home-made color indicator. The colorIndicator is 0.2% Sodium Rhodizonate (this is the saturatedconcentration) and the pH regulating agent is pH˜3.0 by adding citricacid (this pH was optimized by our own experiment). We printed thereagent mixture with a parameter of droplet diameter 500˜600 um, pitch˜1mm and Print speed 10 mm/sec.

For this example, we fabricated a plate, each plate has 48 wells, welldiameter is 3 mm

Center-to-center distance is 6 mm, well height is ˜100 um (controlledusing double-sided tape from Adhesive Research). We then drop 0.7 uL ofsample in each well. Then we cover the well using 175 um thick PET filmand wait for 1 min. Each well is immediately measured after 1 minincubation. For the test, the light source used is the smartphone cameraflash light. And the image is taken using the smartphone's camera.

As assay validation, we calculate 4 key performances: 1. Limit ofDetection (LOD) of each plate; 2. Intra-assay CV % of each plate, 3.Inter-assay CV % of each test day, and 4. Day-to-day CV %. For thisexample we prepared a total of 8 plates, each prepared at a differenttime using different batch of reagent. We perform the test on 2different days and, for each day, we perform the tests on 4 differentplates. On each plates, we perform the assay with 8 differentconcentration from 417 ppb, 213 ppb, 106 ppb, 53.4 ppb, 26.7 ppb, 13.3ppb, 6.7 ppb and 0 ppb. For each concentration, we perform 6 replicates.

FIG. C8 shows the Lead in water test standard curve of individual R, G,B channel. RGB channel signals changes with Pb²⁺ concentration Curve andconverted to a single standard curve using a conversion equationSignal=−0.88*R+G−0.27*B+56.12. The converted data is fitted with 5PLlogistic fitting. Error bar is Standard deviation of 6 replicate wells.The LOD, after conversion is 8.5 ppb.

FIG. C9 shows the sensitivity of all 8 different test plates in thisexample of the invention. Each test plate is prepared separately withdifferent reagent and tested at different time. The average LOD achievedis 8 ppb, which is below the EPA action level at 15 ppb.

FIG. C10 Table of Intra-assay, Inter Assay and Day-to-day CV % of leadin water test. Near LOD, of each tests, the Intra-assay CV %˜4%, theInter-assay CV %˜4% and the Day-to-day CV %˜1.1%

In summary, this example shows a test of lead concentration in tap waterthat shows (1) Sensitivity: average LOD˜8 ppb. All test plates show LODthat meets EPA standard (15 ppb), with the best LOD achieved is 3.9 ppb.(2) Repeatability: Intra-assay CV % at LOD 4%, Inter-assay CV % atLOD˜4% and Day-to-day CV % at LOD˜1.1%

-   D Foodstuff Safety and Allergen Test Using QMAX Device

Another aspect of the present invention provides devices and methods forsafety and allergen test in foodstuff samples.

As summarized above, the devices, systems and methods in the presentinvention may find use in analyzing a foodstuff sample, e.g., a samplefrom raw food, processed food, cooked food, drinking water, etc., forthe presence of foodstuff markers. A foodstuff marker may be anysuitable marker, such as those shown in Table B9, below, that can becaptured by a capturing agent that specifically binds the foodstuffmarker in a CROF device configured with the capturing agent. Theenvironmental sample may be obtained from any suitable source, such astap water, drinking water, prepared food, processed food or raw food,etc. In some embodiments, the presence or absence, or the quantitativelevel of the foodstuff marker in the sample may be indicative of thesafety or harmfulness to a subject if the food stuff is consumed. Insome embodiments, the foodstuff marker is a substance derived from apathogenic or microbial organism that is indicative of the presence ofthe organism in the foodstuff from which the sample was obtained. Insome embodiments, the foodstuff marker is a toxic or harmful substanceif consumed by a subject. In some embodiments, the foodstuff marker is abioactive compound that may unintentionally or unexpectedly alter thephysiology if consumed by the subject. In some embodiments, thefoodstuff marker is indicative of the manner in which the foodstuff wasobtained (grown, procured, caught, harvested, processed, cooked, etc.).In some embodiments, the foodstuff marker is indicative of thenutritional content of the foodstuff. In some embodiments, the foodstuffmarker is an allergen that may induce an allergic reaction if thefoodstuff from which the sample is obtained is consumed by a subject.

In some embodiments, the devices, systems and methods in the presentinvention further includes receiving or providing a report thatindicates the safety or harmfulness for a subject to consume the foodstuff from which the sample was obtained based on information includingthe measured level of the foodstuff marker. The information used toassess the safety of the foodstuff for consumption may include dataother than the type and measured amount of the foodstuff marker. Theseother data may include any health condition associated with the consumer(allergies, pregnancy, chronic or acute diseases, current prescriptionmedications, etc.).

The report may be generated by the device configured to read the CROFdevice, or may be generated at a remote location upon sending the dataincluding the measured amount of the foodstuff marker. In some cases, afood safety expert may be at the remote location or have access to thedata sent to the remote location, and may analyze or review the data togenerate the report. The food safety expert may be a scientist oradministrator at a governmental agency, such as the US Food and DrugAdministration (FDA) or the CDC, a research institution, such as auniversity, or a private company. In certain embodiments, the foodsafety expert may send to the user instructions or recommendations basedon the data transmitted by the device and/or analyzed at the remotelocation.

A list of foodstuff markers is available in Table D1. In someembodiments of the present invention, the QMAX device is used to detectthe presence and/or quantity of analyte, including, but not limited to,the foodstuff markers listed in Table D1.

TABLE D1 Foodstuff Markers Source/Class Marker/target Pathogens/microbesBacillus anthracis (LF), Giardia lamblia, Legionella , Total Coliforms(including fecal coliform and E. Coli), Viruses (enteric) stapylococci(e.g., Staphylococcus epidermidis and Staphylococcus aureus (enterotoxinA, B, C, G, I, cells, TSST-1), Enterrococcus faecalis, Pseudomonasaeruginosa, Escherichia coli (Shiga-like toxin, F4, F5, H, K, O,bacteriophage K1, K5, K13), other gram-positive bacteria, andgram-negative bacilli. Clostridium difficile (Toxin A, B),Bacteroidetes, Cryptosporidium parvum (GP900, p68 or cryptopain,oocyst), Candida albicans, Bacillus anthracis, Bacillusstearothermophilus, Bacillus cereus, Bacillus licheniformis, Bacillussubtilis, Bacillus pumilus, Bacillus badius, Bacillus globigii,Salmonella typhimurium, Escherichia coli O157:H7, Norovirus, Listeriamonocytogenes (internalin), Leptospira interrogans, Leptospira biflexa,Campylobacter jejuni, Campylobacter coli, Clostridium perfringens,Aspergillus flavus (aflatoxins), Aspergillus parasiticus, (aflatoxins),Ebola virus (GP), Histoplasma capsulatum Blastomyces dermatitidis (Aantigen), Gram-positive bacteria (teichoic acid), Gram-ngative bacteria(such as, Pseudomonas aeruginosa, Klebsiella pneumoniae, Salmonellaenteriditis, Enterobacter aerogenes, Enterobacter hermanii, Yersiniaenterocolitica and Shigella sonnei)(LPS), Polio virus, Influenza type Avirus, Disease specific prion (PrP-d), Hepatitis A virus, Toxoplasmagondii, Vibrio cholera, Vibrio parahaemolyticus, Vibrio vulnificus,Enterococcus faecalis, Enterococcus faecium, AngiostrongylusCantonensis, Cyclospora cayetanensis, Entamoeba histolytica, Trichinellaspiralis, Toxins/carcinogens N-methylamino-L-alanine (BMAA), Clostridiumbotulinum neurotoxins, BoNT A, B, Ricin A, B; diphtheria toxin;Aristolochic acid; Colchicine, Ochratoxin A, Sterigmatocystin,Ergotamine, Fumonisins, Fusarin C, domoic acid, Brevetoxin, Mycotoxins,Antimony, Ciguatera fish poisoning, museinol, muscarine, psilocybin,coprius artemetrais, ibotenic acid, amanitin, Nitrite poisoning, Pufferfish (tetrodotoxin), histamine, amnesic, Halogenated Heptachlor,chlordane hydrocarbons Heavy metals Lead, mercury, cadmium, Chromium,Arsenic, Copper, Tin, Zinc, Thallium Allergens peanut (Ara h 1, Ara h 2,Ara h 6), fish, shellfish, mollusks, shrimp (D. pteronyssinustropomyosin allergen, Der p 10) Cod (Gadc1); Atlantic salmon (Sals1);domestic cattle milk (Bosd4, Bosd5, Bosd6, Bosd7, Bosd8); chicken/egg(Gald1, Gald2, Gald3, Gald4, Gald5); shrimp (Mete1); shrimp (Pena1,Peni1); black tiger shrimp (Penm1, Penm2); squid (Todp1), brown gardensnail (Helas1); abalone (Halm1); edible frog (Rane1, Rane2); orientalmustard (Braj1); rapeseed (Bran1); cabbage (Brao3); turnip (Brar1,Brar2); barley (Horv15, Horv16, Horv17, Horv21); rye (Secc20); wheat(Tria18, Tria19, Tria25, Tria26, gliadin); corn (Zeam14, Zeam25); rice(Orys1), celery (Apig1, Apig4, Apig5); carrot (Dauc1, Dauc4); hazelnut(Cora1.04, Cora2, Cora8); strawberry (Fraa1, Fraa3, Fraa4); apple(Mald1, Mald2, Mald3, Mald4); pear (Pyrc1, Pyrc4, Pyrc5); avocado(Persa1); apricot (Pruar1, Pruar3); sweet cherry (Pruav1, Pruav2,Pruav3, Pruav4); European plum (Prud3); almond (Prudu4); peach (Prup3,Prup4); asparagus (Aspao1); saffron crocus (Cros1, Cros2); lettuce(Lacs1); grape (Vitv1); banana (Musxp1); pineapple (Anac1, Anac2); lemon(Citl3); sweet orange (Cits1, Cits2, Cits3); litchi (Litc1); yellowmustard (Sina1); soybean (Glym1, Glym2, Glym3, Glym4); mung bean(Vigr1); peanut (Arah1, Arah2, Arah3, Arah4, Arah5, Arah6, Arah7,Arah8); lentil (Lenc1, Lenc2); pea (Piss1, Piss2); kiwi (Actc1, Actc2);bell pepper (Capa1w, Capa2); tomato (Lyce1, Lyce2, Lyce3); potato(Solat1, Solat2, Solat3, Solat4); Brazil nut (Bere1, Bere2); blackwalnut (Jugn1, Jugn2); English walnut (Jugr1, Jugr2, Jugr3); Cashew(Anao1, Anao2, Anao3); Castor bean (Ricc1); sesame (Sesi1, Sesi2, Sesi3,Sesi4, Sesi5, Sesi6); muskmelon (Cucm1, Cucm2, Cucm3); Chinese-date(Zizm1); Anacardium occidentale (Anao1.0101, Anao1.0102); Apiumgraveolens (Apig1.0101, Apig1.0201); Daucus carota (Dauc1.0101,Dauc1.0102, Dauc1.0103, Dauc1.0104, Dauc1.0105, Dauc1.0201); Citrussinensis (Cits3.0101, Cits3.0102); Glycine max (Glym1.0101, Glym1.0102,Glym3.0101, Glym3.0102); Lens culinaris (Lenc1.0101, Lenc1.0102,Lenc1.0103); Pisum sativum (Piss1.0101, Piss1.0102); Lycopersiconesculentum (Lyce2.0101, Lyce2.0102); Fragaria ananassa (Fraa3.0101,Fraa3.0102, Fraa3.0201, Fraa3.0202, Fraa3.0203, Fraa3.0204, Fraa3.0301);Malus domestica (Mald1.0101, Mald1.0102, Mald1.0103, Mald1.0104,Mald1.0105, Mald1.0106, Mald1.0107, Mald1.0108, Mald1.0109, Mald1.0201,Mald1.0202, Mald1.0203, Mald1.0204, Mald1.0205, Mald1.0206, Mald1.0207,Mald1.0208, Mald1.0301, Mald1.0302, Mald1.0303, Mald1.0304, Mald1.0401,Mald1.0402, Mald1.0403, Mald3.0101w, Mald3.0102w, Mald3.0201w,Mald3.0202w, Mald3.0203w, Mald4.0101, Mald4.0102, Mald4.0201,Mald4.0202, Mald4.0301, Mald4.0302); Prunus avium (Pruav1.0101,Pruav1.0201, Pruav1.0202, Pruav1.0203); and Prunus persica (Prup4.0101,Prup4.0201) Synthetic hormone 17beta-estradiol (E2), estrone (El),estrogen (ES: El + E2 + analogues estradiol (E3)), 17alfa-ethynylestradiol (EE2), 4-nonylphenpol, testosterone,Diethylstilbestrol (DES), recombinant bovine growth hormone (rBGH)Pesticides Dieldrin, carbaryl, chlorpyrifos, parathion, aldrin,endosulfan I, endrin, toxaphene, O-ethyl O-4-nitrophenylphenylphosphono- thioate (EPN), fenitrothion, pirimiphos-methyl,thiabendazole, methiocarb, Carbendazim, deltamethrin, Avermectin,Carbaryl, Cyanazine, Kresoxim, resmethrin, kadethrin, cyhalothrin,biphenthrin, fenpropathrin, allethrin and tralomethrin;aromatic-substituted alkanecarboxylic acid esters such as fenvarerate,flucythrinate, fluvalinate and cycloprothrin; and non-ester compoundssuch as etofenprox, halfenprox (MTI-732), 1-(3-phenoxyphenyl)-4-(4-ethoxyphenyl)-4-methylpentane (MTI-790), 1-(3-phenoxy-4-fluorophenyl)-4-(4-ethoxyphenyl)-4-methylpentane (MTI-800),dimethyl-(4-ethoxyphenyl)-(3-phenoxybenzyloxy)silane (SSI-116),silafluofen and PP-682, carbofuran, triazophos Herbicide atrazine,deethylatrazine, cyanazine, terbuthylazine, terbutryn, molinate,simazine, prometon, promteryn, hydroxyatrazine, 2,6- dichlorobenzamide(BAM), N-dealkylated triazines, mecoprop, thiram, acetochlor, alachlor,Chlorothalonil, Chlorsulfuron, Fenoxaprop ethyl, Linuron, monuron,diuron, Quizalofop-ethyl, Imazalil, Iprodione, Iprovalicarb,Myclobutanil Industrial Dioxin (2,3,7,8-TCDD), 4-tert-octylphenol,bisphenol A (BPA), material/waste Styrene, Di(2-ethylhexyl) phthalate,Dibutyl phthalate (DBP), benzophenone, benzene, trichloroethylene,polychlorinated biphenyl (PCB), nonylphenol, p-cresol, melamine, xylene,Sodium Fluoride Antibiotics 3-Amino-5-morpholinomethyl-2-oxazolidone(AMOZ; tissue bound metabolite of furaltadone), oxytetracycline,rolitetracycline, Actinomycin D, Amikacin sulfate, Aminoglycosides,nitrofuran (AOZ), Chloramphenicol, Doxycycline, Streptomycin,gentamicin, neomycin, kanamycin, sulfamethazine, enrofloxacin,sulfadiazine, enrofloxacin Food coloring/ Tartrazine, ethoxyquin,erythritol, penicillin, Fluoroquinolone, additive/ MalachiteGreen/Leucomalachite Green, C.I. Solvent Yellow 14 preservative (SudanI), Food preparation Acrylamide,2-amino-3-methylimidazo(4,5-f)quinolone, Benzo[a]pyrene Nutritionalcontent Vitamins A (retinol), B12 (cobalmins), B6 (pyridoxine), B1(thiamin), B2 (riboflavin), B3 (niacin), B5 (D-pantothenic acid), B7(biotin), B9 (folic acid), C, D, E (alpha-tocopherol); Other Caffeine,Ovine myofibril proteins, Etodolac

-   E. Uniform Sample Thickness Pressed by an Imprecise Force.

In some embodiments of devices or methods of forming uniform samplethickness by pressing with an imprecise force described herein and inthe provisional 62/456,504, filed on Feb. 8, 2017., which isincorporated herein in the its entirety for all purposes.

In some embodiments, the imprecise force is around 0.01 kg, 0.05 kg, 0.1kg, 0.25 kg, 0.5 kg, 1 kg, 2.5 kg, 5 kg, 7.5 kg, 10 kg, 20 kg, 25 kg, 30kg, 40 kg, 50 kg, 60 kg, 70 kg, 80 kg, 100 kg, 200 kg, or in a rangebetween any two of these values; and a preferred range of 0.5-2 kg, 2-5kg, 5-7.5 kg, 7.5-10 kg, 10-20 kg, 20-40 kg, 40-60 kg, or 60-100 kg.

In some embodiments, the imprecise force is applied by human hand, forexample, e.g., by pinching an object together between a thumb and indexfinger, or by pinching and rubbing an object together between a thumband index finger.

In some embodiments, the hand pressing force is around 0.05 kg, 0.1 kg,0.25 kg, 0.5 kg, 1 kg, 2.5 kg, 5 kg, 7.5 kg, 10 kg, 20 kg, 25 kg, 30 kg,40 kg, 50 kg, 60 kg, or in a range between any two of these values; anda preferred range of 0.5-1 kg, 1-2 kg, 2-4 kg, 4-6 kg, 6-10 kg, 10-20kg, 20-40 kg, or 40-60 kg.

In some embodiments, the hand pressing has a pressure of 0.01 kg/cm²,0.1 kg/cm², 0.5 kg/cm², 1 kg/cm², 2 kg/cm², 2.5 kg/cm², 5 kg/cm², 10kg/cm², 20 kg/cm², 30 kg/cm², 40 kg/cm², 50 kg/cm², 60 kg/cm², 100kg/cm², 150 kg/cm², 200 kg/cm², or a range between any two of thevalues; and a preferred range of 0.1 kg/cm² to 0.5 kg/cm², 0.5 kg/cm² to1 kg/cm², 1 kg/cm² to 5 kg/cm², or 5 kg/cm² to 10 kg/cm².

As used herein, the term “imprecise” in the context of a force (e.g.“imprecise pressing force”) refers to a force that

(a) has a magnitude that is not precisely known or precisely predictableat the time the force is applied;

(b) varies in magnitude from one application of the force to the next;and

(c) the imprecision (i.e. the variation) of the force in (a) and (c) isat least 20% of the total force that actually is applied.

An imprecise force can be applied by human hand, for example, e.g., bypinching an object together between a thumb and index finger, or bypinching and rubbing an object together between a thumb and indexfinger.

-   EA. Imprecise Force, Specify IGS{circumflex over ( )}4/hE-   EA1. A device for forming a thin fluidic sample layer with a uniform    predetermined thickness by pressing with an imprecise pressing    force, comprising:

a first plate, a second plate, and spacers, wherein:

-   -   i. the plates are movable relative to each other into different        configurations;    -   ii. one or both plates are flexible;    -   iii. each of the plates comprises an inner surface that has a        sample contact area for contacting a fluidic sample;    -   iv. each of the plates comprises, on its respective outer        surface, a force area for applying an imprecise pressing force        that forces the plates together;    -   v. one or both of the plates comprise the spacers that are        permanently fixed on the inner surface of a respective plate;    -   vi. the spacers have a predetermined substantially uniform        height that is equal to or less than 200 microns, and a        predetermined fixed inter-spacer-distance;    -   vii. the fourth power of the inter-spacer-distance (IDS) divided        by the thickness (h) and the Young's modulus (E) of the flexible        plate (ISD⁴/(hE)) is 5×10⁶ um³/GPa or less; and    -   viii. at least one of the spacers is inside the sample contact        area;

wherein one of the configurations is an open configuration, in which:the two plates are partially or completely separated apart, the spacingbetween the plates is not regulated by the spacers, and the sample isdeposited on one or both of the plates;

wherein another of the configurations is a closed configuration which isconfigured after the sample is deposited in the open configuration andthe plates are forced to the closed configuration by applying theimprecise pressing force on the force area; and in the closedconfiguration: at least part of the sample is compressed by the twoplates into a layer of highly uniform thickness and is substantiallystagnant relative to the plates, wherein the uniform thickness of thelayer is confined by the sample contact areas of the two plates and isregulated by the plates and the spacers.

-   EA2. A method of forming a thin fluidic sample layer with a uniform    predetermined thickness by pressing with an imprecise pressing    force, comprising the steps of:    -   (a) obtaining a first plate, a second plate, and spacers,        wherein:        -   i. the plates are movable relative to each other into            different configurations;        -   ii. one or both plates are flexible;        -   iii. each of the plates comprises an inner surface that has            a sample contact area for contacting a fluidic sample;        -   iv. each of the plates comprises, on its respective outer            surface, a force area for applying an imprecise pressing            force that forces the plates together;        -   v. one or both of the plates comprise the spacers that are            permanently fixed on the inner surface of a respective            plate;        -   vi. the spacers have a predetermined substantially uniform            height that is equal to or less than 200 microns, and a            predetermined fixed inter-spacer-distance;        -   vii. the fourth power of the inter-spacer-distance (IDS)            divided by the thickness (h) and the Young's modulus (E) of            the flexible plate (ISD⁴/(hE)) is 5×10⁶ um³/GPa or less; and        -   viii. at least one of the spacers is inside the sample            contact area;    -   (b) obtaining a fluidic sample;    -   (c) depositing the sample on one or both of the plates; when the        plates are configured in an open configuration, wherein the open        configuration is a configuration in which the two plates are        partially or completely separated apart and the spacing between        the plates is not regulated by the spacers;    -   (d) after (c), using the two plates to compress at least part of        the sample into a layer of substantially uniform thickness that        is confined by the sample contact surfaces of the plates,        wherein the uniform thickness of the layer is regulated by the        spacers and the plates, wherein the compressing comprises:        -   bringing the two plates together; and        -   conformable pressing, either in parallel or sequentially, an            area of at least one of the plates to press the plates            together to a closed configuration, wherein the conformable            pressing generates a substantially uniform pressure on the            plates over the at least part of the sample, and the            pressing spreads the at least part of the sample laterally            between the sample contact surfaces of the plates, and            wherein the closed configuration is a configuration in which            the spacing between the plates in the layer of uniform            thickness region is regulated by the spacers; and wherein            the reduced thickness of the sample reduces the time for            mixing the reagents on the storage site with the sample, and        -   wherein the force that presses the two plates into the            closed configuration is an imprecise pressing force provided            by human hand.-   EB. Hand pressing, Specify Spacer Hardness-Contact Area Product-   EB1. A device for forming a thin fluidic sample layer with a uniform    predetermined thickness by pressing with an imprecise force,    comprising:

a first plate, a second plate, and spacers, wherein:

-   -   i. the plates are movable relative to each other into different        configurations;    -   ii. one or both plates are flexible;    -   iii. each of the plates comprises, on its respective inner        surface, a sample contact area for contacting and/or compressing        a fluidic sample;    -   iv. each of the plates comprises, on its respective outer        surface, an area for applying a force that forces the plates        together;    -   v. one or both of the plates comprise the spacers that are        permanently fixed on the inner surface of a respective plate;    -   vi. the spacers have a predetermined substantially uniform        height that is equal to or less than 200 microns, a        predetermined width, and a predetermined inter-spacer-distance;    -   vii. a ratio of the inter-spacer-distance to the spacer width is        1.5 or larger; and    -   viii. at least one of the spacers is inside the sample contact        area;

wherein one of the configurations is an open configuration, in which:the two plates are partially or completely separated apart, the spacingbetween the plates is not regulated by the spacers, and the sample isdeposited on one or both of the plates;

wherein another of the configurations is a closed configuration which isconfigured after the sample deposition in the open configuration; and inthe closed configuration: at least part of the sample is compressed bythe two plates into a layer of highly uniform thickness and issubstantially stagnant relative to the plates, wherein the uniformthickness of the layer is confined by the sample contact areas of thetwo plates and is regulated by the plates and the spacers; and

wherein the force that presses the two plates into the closedconfiguration is an imprecise pressing force provided by human hand.

-   EB2. A method of forming a thin fluidic sample layer with a uniform    predetermined thickness by pressing with an imprecise pressing    force, comprising the steps of:    -   (a) obtaining a first plate, a second plate, and spacers,        wherein:        -   i. the plates are movable relative to each other into            different configurations;        -   ii. one or both plates are flexible;        -   iii. each of the plates comprises, on its respective inner            surface, a sample contact area for contacting and/or            compressing a fluidic sample;        -   iv. each of the plates comprises, on its respective outer            surface, an area for applying a force that forces the plates            together;        -   v. one or both of the plates comprise the spacers that are            permanently fixed on the inner surface of a respective            plate;        -   vi. the spacers have a predetermined substantially uniform            height that is equal to or less than 200 microns, a            predetermined width, and a predetermined            inter-spacer-distance;        -   vii. a ratio of the inter-spacer-distance to the spacer            width is 1.5 or larger; and        -   viii. at least one of the spacers is inside the sample            contact area;    -   (b) obtaining a fluidic sample;    -   (c) depositing the sample on one or both of the plates; when the        plates are configured in an open configuration, wherein the open        configuration is a configuration in which the two plates are        partially or completely separated apart and the spacing between        the plates is not regulated by the spacers;    -   (d) after (c), using the two plates to compress at least part of        the sample into a layer of substantially uniform thickness that        is confined by the sample contact surfaces of the plates,        wherein the uniform thickness of the layer is regulated by the        spacers and the plates, wherein the compressing comprises:        -   bringing the two plates together; and        -   conformable pressing, either in parallel or sequentially, an            area of at least one of the plates to press the plates            together to a closed configuration, wherein the conformable            pressing generates a substantially uniform pressure on the            plates over the at least part of the sample, and the            pressing spreads the at least part of the sample laterally            between the sample contact surfaces of the plates, and            wherein the closed configuration is a configuration in which            the spacing between the plates in the layer of uniform            thickness region is regulated by the spacers; and wherein            the reduced thickness of the sample reduces the time for            mixing the reagents on the storage site with the sample, and        -   wherein the force that presses the two plates into the            closed configuration is an imprecise pressing force provided            by human hand.-   EC. Hand Pressing, Specify IDS/hE & Spacer Hardness-Contact Area    Product-   EC1. A device for forming a thin fluidic sample layer with a uniform    predetermined thickness by pressing with an imprecise force,    comprising:    -   a first plate, a second plate, and spacers, wherein:        -   i. the plates are movable relative to each other into            different configurations;        -   ii. one or both plates are flexible;        -   iii. each of the plates comprises, on its respective inner            surface, a sample contact area for contacting and/or            compressing a fluidic sample;        -   iv. each of the plates comprises, on its respective outer            surface, an area for applying a force that forces the plates            together;        -   v. one or both of the plates comprise the spacers that are            permanently fixed on the inner surface of a respective            plate;        -   vi. the spacers have a predetermined substantially uniform            height that is equal to or less than 200 microns, a            predetermined width, and a predetermined            inter-spacer-distance;        -   vii. a ratio of the inter-spacer-distance to the spacer            width is 1.5 or larger; and        -   viii. at least one of the spacers is inside the sample            contact area;

wherein one of the configurations is an open configuration, in which:the two plates are partially or completely separated apart, the spacingbetween the plates is not regulated by the spacers, and the sample isdeposited on one or both of the plates;

wherein another of the configurations is a closed configuration which isconfigured after the sample deposition in the open configuration; and inthe closed configuration: at least part of the sample is compressed bythe two plates into a layer of highly uniform thickness and issubstantially stagnant relative to the plates, wherein the uniformthickness of the layer is confined by the sample contact areas of thetwo plates and is regulated by the plates and the spacers;

wherein the force that presses the two plates into the closedconfiguration is imprecise, and is provided by human hand.

-   EC2. A method of forming a thin fluidic sample layer with a uniform    predetermined thickness by pressing with an imprecise pressing    force, comprising the steps of:    -   (a) obtaining a first plate, a second plate, and spacers,        wherein:        -   i. the plates are movable relative to each other into            different configurations;        -   ii. one or both plates are flexible;        -   iii. each of the plates comprises, on its respective inner            surface, a sample contact area for contacting and/or            compressing a fluidic sample;        -   iv. each of the plates comprises, on its respective outer            surface, an area for applying a force that forces the plates            together;        -   v. one or both of the plates comprise the spacers that are            permanently fixed on the inner surface of a respective            plate;        -   vi. the spacers have a predetermined substantially uniform            height that is equal to or less than 200 microns, a            predetermined width, and a predetermined            inter-spacer-distance;        -   vii. a ratio of the inter-spacer-distance to the spacer            width is 1.5 or larger; and        -   viii. at least one of the spacers is inside the sample            contact area;    -   (b) obtaining a fluidic sample;    -   (c) depositing the sample on one or both of the plates; when the        plates are configured in an open configuration, wherein the open        configuration is a configuration in which the two plates are        partially or completely separated apart and the spacing between        the plates is not regulated by the spacers;    -   (d) after (c), using the two plates to compress at least part of        the sample into a layer of substantially uniform thickness that        is confined by the sample contact surfaces of the plates,        wherein the uniform thickness of the layer is regulated by the        spacers and the plates, wherein the compressing comprises:        -   bringing the two plates together; and        -   conformable pressing, either in parallel or sequentially, an            area of at least one of the plates to press the plates            together to a closed configuration, wherein the conformable            pressing generates a substantially uniform pressure on the            plates over the at least part of the sample, and the            pressing spreads the at least part of the sample laterally            between the sample contact surfaces of the plates, and            wherein the closed configuration is a configuration in which            the spacing between the plates in the layer of uniform            thickness region is regulated by the spacers; and wherein            the reduced thickness of the sample reduces the time for            mixing the reagents on the storage site with the sample, and        -   wherein the force that presses the two plates into the            closed configuration is an imprecise pressing force provided            by human hand.-   ED. Hand Pressing, Specify Pillar Spacer and Ratio of IDS/W-   ED 1. A device for forming a thin fluidic sample layer with a    uniform predetermined thickness by pressing with an imprecise force,    comprising:    -   a first plate, a second plate, and spacers, wherein:        -   i. the plates are movable relative to each other into            different configurations;        -   ii. one or both plates are flexible;        -   iii. each of the plates comprises, on its respective inner            surface, a sample contact area for contacting and/or            compressing a fluidic sample;        -   iv. each of the plates comprises, on its respective outer            surface, an area for applying a force that forces the plates            together;        -   v. one or both of the plates comprise the spacers that are            permanently fixed on the inner surface of a respective            plate;        -   vi. the spacers have a predetermined substantially uniform            height that is equal to or less than 200 microns, a            predetermined width, and a predetermined            inter-spacer-distance;        -   vii. a ratio of the inter-spacer-distance to the spacer            width is 1.5 or larger.        -   viii. at least one of the spacers is inside the sample            contact area; and

wherein one of the configurations is an open configuration, in which:the two plates are partially or completely separated apart, the spacingbetween the plates is not regulated by the spacers, and the sample isdeposited on one or both of the plates;

wherein another of the configurations is a closed configuration which isconfigured after the sample deposition in the open configuration; and inthe closed configuration: at least part of the sample is compressed bythe two plates into a layer of highly uniform thickness and issubstantially stagnant relative to the plates, wherein the uniformthickness of the layer is confined by the sample contact areas of thetwo plates and is regulated by the plates and the spacers;

wherein the force that presses the two plates into the closedconfiguration is imprecise, and is provided by human hand.

-   ED2. A method of forming a thin fluidic sample layer with a uniform    predetermined thickness by pressing with an imprecise pressing    force, comprising the steps of:    -   (a) obtaining a first plate, a second plate, and spacers,        wherein:        -   i. the plates are movable relative to each other into            different configurations;        -   ii. one or both plates are flexible;        -   iii. each of the plates comprises, on its respective inner            surface, a sample contact area for contacting and/or            compressing a fluidic sample;        -   iv. each of the plates comprises, on its respective outer            surface, an area for applying a force that forces the plates            together;        -   v. one or both of the plates comprise the spacers that are            permanently fixed on the inner surface of a respective            plate;        -   vi. the spacers have a predetermined substantially uniform            height that is equal to or less than 200 microns, a            predetermined width, and a predetermined            inter-spacer-distance;        -   vii. a ratio of the inter-spacer-distance to the spacer            width is 1.5 or larger.        -   viii. at least one of the spacers is inside the sample            contact area; and    -   (b) obtaining a fluidic sample;    -   (c) depositing the sample on one or both of the plates; when the        plates are configured in an open configuration, wherein the open        configuration is a configuration in which the two plates are        partially or completely separated apart and the spacing between        the plates is not regulated by the spacers;    -   (d) after (c), using the two plates to compress at least part of        the sample into a layer of substantially uniform thickness that        is confined by the sample contact surfaces of the plates,        wherein the uniform thickness of the layer is regulated by the        spacers and the plates, wherein the compressing comprises:        -   bringing the two plates together; and        -   conformable pressing, either in parallel or sequentially, an            area of at least one of the plates to press the plates            together to a closed configuration, wherein the conformable            pressing generates a substantially uniform pressure on the            plates over the at least part of the sample, and the            pressing spreads the at least part of the sample laterally            between the sample contact surfaces of the plates, and            wherein the closed configuration is a configuration in which            the spacing between the plates in the layer of uniform            thickness region is regulated by the spacers; and wherein            the reduced thickness of the sample reduces the time for            mixing the reagents on the storage site with the sample, and            wherein the force that presses the two plates into the            closed configuration is an imprecise pressing force provided            by human hand.-   EE. Volume Determination, Specify IGS{circumflex over ( )}4/hE-   EE1. A device for determining a relevant sample volume by pressing    with an imprecise force provided by human hand, comprising:    -   a first plate, a second plate, spacers, and an        area-determination device, wherein:        -   i. the plates are movable relative to each other into            different configurations;        -   ii. one or both plates are flexible;        -   iii. each of the plates comprises, on its respective inner            surface, a sample contact area for contacting and/or            compressing a fluidic sample that has a relevant volume to            be measured;        -   iv. each of the plates comprises, on its respective outer            surface, an area for applying a force that forces the plates            together;        -   v. one or both of the plates comprise the spacers that are            permanently fixed on the inner surface of a respective            plate;        -   vi. the spacers have a predetermined substantially uniform            height that is equal to or less than 200 microns, and a            predetermined constant inter-spacer-distance;        -   vii. a fourth power of the inter-spacer-distance (IDS)            divided by the thickness (h) and the Young's modulus (E) of            the flexible plate (ISD⁴/(hE)) is 5×10⁶ um³/GPa or less.        -   viii. at least one of the spacers is inside the sample            contact area; and        -   ix. the area-determination device is configured to determine            the lateral area of the relevant volume;

wherein one of the configurations is an open configuration, in which:the two plates are partially or completely separated apart, the spacingbetween the plates is not regulated by the spacers, and the sample isdeposited on one or both of the plates;

wherein another of the configurations is a closed configuration which isconfigured after the sample deposition in the open configuration; and inthe closed configuration: at least part of the sample is compressed bythe two plates into a layer of highly uniform thickness and issubstantially stagnant relative to the plates, wherein the uniformthickness of the layer is confined by the sample contact areas of thetwo plates and is regulated by the plates and the spacers;

wherein the relevant volume of the sample is a partial or entire volumeof the uniform thickness layer and the value of the relevant volume isdetermined by the uniform thickness and the determined lateral area; and

wherein the force that presses the two plates into the closedconfiguration is imprecise, and is provided by human hand.

The device of any prior embodiment, wherein the area-determinationdevice is a camera.

-   -   The area-determination device comprises an area in the sample        contact area of a plate, wherein the area is less than 1/100,        1/20, 1/10, ⅙, ⅕, ¼, ⅓, ½, ⅔ of the sample contact area, or in a        range between any of the two values.    -   The area-determination device comprises a camera and an area in        the sample contact area of a plate, wherein the area is in        contact with the sample.

-   EE2. A method of forming a thin fluidic sample layer with a uniform    predetermined thickness by pressing with an imprecise pressing    force, comprising the steps of:    -   (a) obtaining a first plate, a second plate, and spacers,        wherein:        -   i. the plates are movable relative to each other into            different configurations;        -   ii. one or both plates are flexible;        -   iii. each of the plates comprises, on its respective inner            surface, a sample contact area for contacting and/or            compressing a fluidic sample that has a relevant volume to            be measured;        -   iv. each of the plates comprises, on its respective outer            surface, an area for applying a force that forces the plates            together;        -   v. one or both of the plates comprise the spacers that are            permanently fixed on the inner surface of a respective            plate;        -   vi. the spacers have a predetermined substantially uniform            height that is equal to or less than 200 microns, and a            predetermined constant inter-spacer-distance;        -   vii. a fourth power of the inter-spacer-distance (IDS)            divided by the thickness (h) and the Young's modulus (E) of            the flexible plate (ISD⁴/(hE)) is 5×10⁶ um³/GPa or less.        -   viii. at least one of the spacers is inside the sample            contact area; and        -   ix. the area-determination device is configured to determine            the lateral area of the relevant volume;    -   (b) obtaining a fluidic sample;    -   (c) depositing the sample on one or both of the plates; when the        plates are configured in an open configuration, wherein the open        configuration is a configuration in which the two plates are        partially or completely separated apart and the spacing between        the plates is not regulated by the spacers;    -   (d) after (c), using the two plates to compress at least part of        the sample into a layer of substantially uniform thickness that        is confined by the sample contact surfaces of the plates,        wherein the uniform thickness of the layer is regulated by the        spacers and the plates, wherein the compressing comprises:        -   bringing the two plates together; and        -   conformable pressing, either in parallel or sequentially, an            area of at least one of the plates to press the plates            together to a closed configuration, wherein the conformable            pressing generates a substantially uniform pressure on the            plates over the at least part of the sample, and the            pressing spreads the at least part of the sample laterally            between the sample contact surfaces of the plates, and            wherein the closed configuration is a configuration in which            the spacing between the plates in the layer of uniform            thickness region is regulated by the spacers; and wherein            the reduced thickness of the sample reduces the time for            mixing the reagents on the storage site with the sample, and        -   wherein the force that presses the two plates into the            closed configuration is an imprecise pressing force provided            by human hand.

-   EF. Volume Determination, Specify IGS{circumflex over ( )}4/hE

-   EF1. A device for determining a relevant sample volume by pressing    with an imprecise force provided by human hand, comprising:    -   a first plate, a second plate, spacers, and area-determination        device, wherein:        -   i. the plates are movable relative to each other into            different configurations;        -   ii. one or both plates are flexible;        -   iii. each of the plates comprises, on its respective inner            surface, a sample contact area for contacting and/or            compressing a fluidic sample that has a relevant volume to            be measured;        -   iv. each of the plates comprises, on its respective outer            surface, an area for applying a force that forces the plates            together;        -   v. one or both of the plates comprise the spacers that are            permanently fixed on the inner surface of a respective            plate;        -   vi. the spacers have a predetermined substantially uniform            height that is equal to or less than 200 microns, and a            predetermined constant inter-spacer-distance;        -   vii. a fourth power of the inter-spacer-distance (IDS)            divided by the thickness (h) and the Young's modulus (E) of            the flexible plate (ISD⁴/(hE)) is 5×10⁶ um³/GPa or less.        -   viii. at least one of the spacers is inside the sample            contact area; and        -   ix. the area-determination device is configured to determine            the lateral area of the relevant volume;

wherein one of the configurations is an open configuration, in which:the two plates are partially or completely separated apart, the spacingbetween the plates is not regulated by the spacers, and the sample isdeposited on one or both of the plates;

wherein another of the configurations is a closed configuration which isconfigured after the sample deposition in the open configuration; and inthe closed configuration: at least part of the sample is compressed bythe two plates into a layer of highly uniform thickness and issubstantially stagnant relative to the plates, wherein the uniformthickness of the layer is confined by the sample contact areas of thetwo plates and is regulated by the plates and the spacers;

wherein the relevant volume of the sample is a partial or entire volumeof the uniform thickness layer and the value of the relevant volume isdetermined by the uniform thickness and the determined lateral area; and

wherein the force that presses the two plates into the closedconfiguration is imprecise, and is provided by human hand.

-   EF2. A method of forming a thin fluidic sample layer with a uniform    predetermined thickness by pressing with an imprecise pressing    force, comprising the steps of:    -   (a) obtaining a first plate, a second plate, and spacers,        wherein:        -   i. the plates are movable relative to each other into            different configurations;        -   ii. one or both plates are flexible;        -   iii. each of the plates comprises, on its respective inner            surface, a sample contact area for contacting and/or            compressing a fluidic sample that has a relevant volume to            be measured;        -   iv. each of the plates comprises, on its respective outer            surface, an area for applying a force that forces the plates            together;        -   v. one or both of the plates comprise the spacers that are            permanently fixed on the inner surface of a respective            plate;        -   vi. the spacers have a predetermined substantially uniform            height that is equal to or less than 200 microns, and a            predetermined constant inter-spacer-distance;        -   vii. a fourth power of the inter-spacer-distance (IDS)            divided by the thickness (h) and the Young's modulus (E) of            the flexible plate (ISD⁴/(hE)) is 5×10⁶ um³/GPa or less.        -   viii. at least one of the spacers is inside the sample            contact area; and        -   ix. the area-determination device is configured to determine            the lateral area of the relevant volume;    -   (b) obtaining a fluidic sample;    -   (c) depositing the sample on one or both of the plates; when the        plates are configured in an open configuration, wherein the open        configuration is a configuration in which the two plates are        partially or completely separated apart and the spacing between        the plates is not regulated by the spacers;    -   (d) after (c), using the two plates to compress at least part of        the sample into a layer of substantially uniform thickness that        is confined by the sample contact surfaces of the plates,        wherein the uniform thickness of the layer is regulated by the        spacers and the plates, wherein the compressing comprises:        -   bringing the two plates together; and        -   conformable pressing, either in parallel or sequentially, an            area of at least one of the plates to press the plates            together to a closed configuration, wherein the conformable            pressing generates a substantially uniform pressure on the            plates over the at least part of the sample, and the            pressing spreads the at least part of the sample laterally            between the sample contact surfaces of the plates, and            wherein the closed configuration is a configuration in which            the spacing between the plates in the layer of uniform            thickness region is regulated by the spacers; and wherein            the reduced thickness of the sample reduces the time for            mixing the reagents on the storage site with the sample, and        -   wherein the force that presses the two plates into the            closed configuration is an imprecise pressing force provided            by human hand.-   EG. More Embodiments

The term “imprecise force” refers to a force that has a magnitude thatis completely unknown, known only in a magnitude range but not in aparticular magnitude value (the magnitude range varies at least 20% fromthe minimum to the maximum of the range), or unpredictable at the timethat a force is applied. Examples of an imprecise force include that themagnitude of an imprecise force may vary from one application of theforce to the next, may be uneven across the area upon which the force isapplied, and may vary over the time that the force is being applied. Animprecise force does not need to be measured at the time that it isapplied.

The devices or methods of any prior embodiment, wherein the deformablesample is a fluidic sample.

The devices or methods of any prior embodiment, wherein the deformablesample is a liquid sample.

The devices or methods of any prior embodiment, wherein the imprecisionforce has a variation at least 30% of the total force that actually isapplied.

The devices or methods of any prior embodiment, wherein the imprecisionforce has a variation at least 20%, 30%, 40%, 50%, 60, 70%, 80%, 90%100%, 150%, 200%, 300%, 500%, or in a range of any two values, of thetotal force that actually is applied.

-   1. The device of any prior embodiment, wherein spacers have a flat    top.-   2. The device of any prior embodiment, wherein the device is further    configured to have, after the pressing force is removed, a sample    thickness that is substantially the same in thickness and uniformity    as that when the force is applied.-   3. The device of any prior embodiment, wherein the imprecise force    is provided by human hand.-   4. The device of any prior embodiment, wherein the inter spacer    distance is substantially constant.-   5. The device of any prior embodiment, wherein the inter spacer    distance is substantially periodic in the area of the uniform sample    thickness area.-   6. The device of any prior embodiment, wherein the multiplication    product of the filling factor and the Young's modulus of the spacer    is 2 MPa or larger.-   7. The device of any prior embodiment, wherein the force is applied    by hand directly or indirectly.-   8. The device of any prior embodiment, wherein the force applied is    in the range of 5 N to 20 N.-   9. The device of any prior embodiment wherein the highly uniform    layer has a thickness that varies by less than 15%, 10%, or 5% of an    average thickness.-   10. The device of any prior embodiment, wherein the imprecise force    is applied by pinching the device between a thumb and forefinger.-   11. The device of any prior embodiment, wherein the predetermined    sample thickness is larger than the spacer height.-   12. The device of any prior embodiment, wherein the device holds    itself in the closed configuration after the pressing force has been    removed.-   13. The device of any prior embodiment, wherein the uniform    thickness sample layer area is larger than that area upon which the    pressing force is applied.-   14. The device of any prior embodiment, wherein the spacers do not    significantly deform during application of the pressing force.-   15. The device of any prior embodiment, wherein the pressing force    is not predetermined beforehand and is not measured.-   F. Binding Site and Storage Site on the Same Plate

Another aspect of the present invention provides devices and methods forbio/chemical assays using QMAX device in which binding site and storagesite are on the same plate, meaning both capture agent and second agentare coated on the same plate.

-   FA1. A method for assaying a sample, comprising    -   (a) obtaining a first plate comprising, on its inner surface, a        sample contact area for contacting a sample that contains a        target analyte;    -   (b) obtaining a second plate comprising a sample contact area        that comprises an assaying area, wherein the assaying area        comprises        -   (i) an immobilized capture agent that binds a target analyte            in a sample, and        -   (ii) a second agent that is capable of, upon contacting the            sample, diffusing in the sample;        -   wherein the first plate and second plate are movable            relative to each other into different configurations,            including an open and a closed configurations;    -   (c) depositing, in the open configuration, the sample on one or        both of the sample contact areas of the plates, wherein in the        open configuration, the sample contact areas of the plates are        separated larger than 200 um;    -   (d) after (c), bringing the two plates to a closed        configuration, wherein, in the closed configuration, at least        part of the sample deposited in (c) is confined between the        sample contact areas of the two plates, and has an average        thickness in the range of 0.01 to 200 μm; and    -   (e) detecting a signal related to an analyte that is captured by        the binding site.-   FB1. A device for performing a competitive assay, comprising:    -   a first plate comprising, on its inner surface, a sample contact        area for contacting a sample that contains a target analyte;    -   a second plate comprising a sample contact area that comprises        an assaying area,        -   wherein the assaying area comprises        -   (i) an immobilized capture agent that binds a target analyte            in a sample, and        -   (ii) a second agent that is capable of, upon contacting the            sample, diffusing in the sample;        -   wherein the first plate and second plate are movable            relative to each other into different configurations;        -   wherein one of the configurations is an open configuration,            in which the plates are partially or entirely separated            apart, and the average spacing between the sample contact            areas of the plates is larger than 300 um; and        -   wherein another configuration is a closed configuration in            which the average spacing between the sample contact areas            of the plates is 200 μm or less.

The method or device of any prior embodiment, wherein the capture agentsand the second agents are separated by a distance that is at least 2times less than the average spacing between the sample contact area ofthe two plates.

The method or device of any prior embodiment, wherein the capture agentsand the second agents are separated by a distance that is at least 2times, 3 times, 5 times, 10 times, 20 times, 30 times, 50 times, 100times, 200 times,300 times,500 times, 1000 times, 2000 times, 5000times, 10000 times, 5000 times, less than the average spacing betweenthe sample contact area of the two plates, or in a range of any twovalues.

The method or device of any prior embodiment, wherein the signal relatedto the analyte captured by the capture agent are the signals coming from(i) the analyte captured by the capture agent, (ii) the label attachedan analyte that is captured by the binding site, or (iii) both (i) and(ii).

The method or device of any prior embodiment, wherein one or both of thesample contact areas comprise spacers, wherein the spacers regulate thespacing between the sample contact areas of the plates when the platesare in the closed configuration.

The method of any prior embodiment, wherein the spacing between thesample contact areas when the plates are in a closed configuration isregulated by spacers.

The device of any prior embodiment, wherein the device further comprisesspacers that regulate the spacing between the sample contact areas whenthe plates are in a closed configuration.

The method or device of any prior embodiment, wherein the storage sitefurther comprises another reagent.

The method or device of any prior embodiment, wherein the binding sitecomprises, in addition to immobilized capture agent, another reagentthat is, upon contacting the sample, capable of diffusion in the sample,

The method or device of any prior embodiment, wherein the detection ofthe signal is electrical, optical, or both. (Will add more on thedetection later. Fluorescence, SPR, etc.).

The method or device of any prior embodiment, wherein the sample is ablood sample (whole blood, plasma, or serum).

The method or device of any prior embodiment, wherein the material offluorescent microsphere is dielectric, (e.g. SiO2, Polystyrene,) or thecombination of dielectric materials thereof.

The method or device of any prior embodiment, which comprises steps ofadding the detection agent of said fluorescence label to the first plateto bind competitive agent.

The method or device of any prior embodiment, which comprises steps ofwashing after the detection agent is added.

The embodiments in these applications herein incorporated can beregarded in combination with one another or as a single invention,rather than as discrete and independent filings.

Moreover, the exemplary assay recipes disclosed herein are applicable toembodiments including but not limited to: bio/chemical assays, QMAXcards and systems, QMAX with hinges, notches, recessed edges andsliders, assays and devices with uniform sample thickness, smartphonedetection systems, cloud computing designs, various detection methods,labels, capture agents and detection agents, analytes, diseases,applications, and samples; the various embodiments are disclosed,described, and/or referred to in the aforementioned applications, all ofwhich are hereby incorporated in reference by their entireties.

-   Other Embodiments

The present invention includes a variety of embodiments, which can becombined in multiple ways as long as the various components do notcontradict one another. The embodiments should be regarded as a singleinvention file: each filing has other filing as the references and isalso referenced in its entirety and for all purpose, rather than as adiscrete independent. These embodiments include not only the disclosuresin the current file, but also the documents that are herein referenced,incorporated, or to which priority is claimed.

-   (1) Definitions

The terms used in describing the devices, systems, and methods hereindisclosed are defined in the current application, or in PCT Application(designating U.S.) Nos. PCT/US2016/045437 and PCT/US0216/051775, whichwere respectively filed on Aug. 10, 2016 and Sep. 14, 2016, U.S.Provisional Application No. 62/456,065, which was filed on Feb. 7, 2017,U.S. Provisional Application No. 62/456,287, which was filed on Feb. 8,2017, and U.S. Provisional Application No. 62/456,504, which was filedon Feb. 8, 2017, all of which applications are incorporated herein intheir entireties for all purposes.

The terms “CROF Card (or card)”, “COF Card”, “QMAX-Card”, “Q-Card”,“CROF device”, “COF device”, “QMAX-device”, “CROF plates”, “COF plates”,and “QMAX-plates” are interchangeable, except that in some embodiments,the COF card does not comprise spacers; and the terms refer to a devicethat comprises a first plate and a second plate that are movablerelative to each other into different configurations (including an openconfiguration and a closed configuration), and that comprises spacers(except some embodiments of the COF card) that regulate the spacingbetween the plates. The term “X-plate” refers to one of the two platesin a CROF card, wherein the spacers are fixed to this plate. Moredescriptions of the COF Card, CROF Card, and X-plate are given in theprovisional application serial nos. 62/456,065, filed on Feb. 7, 2017,which is incorporated herein in its entirety for all purposes.

-   (2) Q-Card, Spacer and Uniform Sample Thickness

The devices, systems, and methods herein disclosed can include or useQ-cards, spacers, and uniform sample thickness embodiments for sampledetection, analysis, and quantification. In some embodiments, the Q-cardcomprises spacers, which help to render at least part of the sample intoa layer of high uniformity. The structure, material, function, variationand dimension of the spacers, as well as the uniformity of the spacersand the sample layer, are herein disclosed, or listed, described, andsummarized in PCT Application (designating U.S.) Nos. PCT/US2016/045437and PCT/US0216/051775, which were respectively filed on Aug. 10, 2016and Sep. 14, 2016, US Provisional Application No. 62/456,065, which wasfiled on Feb. 7, 2017, U.S. Provisional Application No. 62/456,287,which was filed on Feb. 8, 2017, and U.S. Provisional Application No.62/456,504, which was filed on Feb. 8, 2017, all of which applicationsare incorporated herein in their entireties for all purposes.

-   (3) Hinges, Opening Notches, Recessed Edge and Sliders

The devices, systems, and methods herein disclosed can include or useQ-cards for sample detection, analysis, and quantification. In someembodiments, the Q-card comprises hinges, notches, recesses, andsliders, which help to facilitate the manipulation of the Q card and themeasurement of the samples. The structure, material, function, variationand dimension of the hinges, notches, recesses, and sliders are hereindisclosed, or listed, described, and summarized in PCT Application(designating U.S.) Nos. PCT/US2016/045437 and PCT/US0216/051775, whichwere respectively filed on Aug. 10, 2016 and Sep. 14, 2016, U.S.Provisional Application No. 62/456,065, which was filed on Feb. 7, 2017,U.S. Provisional Application No. 62/456,287, which was filed on Feb. 8,2017, and U.S. Provisional Application No. 62/456,504, which was filedon Feb. 8, 2017, all of which applications are incorporated herein intheir entireties for all purposes.

In some embodiments of QMAX, the sample contact area of one or both ofthe plates comprises a compressed open flow monitoring surfacestructures (MSS) that are configured to monitoring how much flow hasoccurred after COF. For examples, the MSS comprises, in someembodiments, shallow square array, which will cause friction to thecomponents (e.g. blood cells in a blood) in a sample. By checking thedistributions of some components of a sample, one can obtain informationrelated to a flow, under a COF, of the sample and its components.

The depth of the MSS can be 1/1000, 1/100, 1/100, ⅕, ½ of the spacerheight or in a range of any two values, and in either protrusion or wellform.

-   (4) Q-Card, Sliders, and Smartphone Detection System

The devices, systems, and methods herein disclosed can include or useQ-cards for sample detection, analysis, and quantification. In someembodiments, the Q-cards are used together with sliders that allow thecard to be read by a smartphone detection system. The structure,material, function, variation, dimension and connection of the Q-card,the sliders, and the smartphone detection system are herein disclosed,or listed, described, and summarized in PCT Application (designatingU.S.) Nos. PCT/US2016/045437 and PCT/US0216/051775, which wererespectively filed on Aug. 10, 2016 and Sep. 14, 2016, U.S. ProvisionalApplication No. 62/456,065, which was filed on Feb. 7, 2017, U.S.Provisional Application No. 62/456,287, which was filed on Feb. 8, 2017,and U.S. Provisional Application No. 62/456,504, which was filed on Feb.8, 2017, all of which applications are incorporated herein in theirentireties for all purposes.

-   (5) Detection Methods

The devices, systems, and methods herein disclosed can include or beused in various types of detection methods. The detection methods areherein disclosed, or listed, described, and summarized in PCTApplication (designating U.S.) Nos. PCT/US2016/045437 andPCT/US0216/051775, which were respectively filed on Aug. 10, 2016 andSep. 14, 2016, U.S. Provisional Application No. 62/456,065, which wasfiled on Feb. 7, 2017, U.S. Provisional Application No. 62/456,287,which was filed on Feb. 8, 2017, and U.S. Provisional Application No.62/456,504, which was filed on Feb. 8, 2017, all of which applicationsare incorporated herein in their entireties for all purposes.

-   (6) Labels, Capture Agent and Detection Agent

The devices, systems, and methods herein disclosed can employ varioustypes of labels, capture agents, and detection agents that are used foranalytes detection. The labels are herein disclosed, or listed,described, and summarized in PCT Application (designating U.S.) Nos.PCT/US2016/045437 and PCT/US0216/051775, which were respectively filedon Aug. 10, 2016 and Sep. 14, 2016, U.S. Provisional Application No.62/456,065, which was filed on Feb. 7, 2017, U.S. ProvisionalApplication No. 62/456,287, which was filed on Feb. 8, 2017, and U.S.Provisional Application No. 62/456,504, which was filed on Feb. 8, 2017,all of which applications are incorporated herein in their entiretiesfor all purposes.

-   (7) Analytes

The devices, systems, and methods herein disclosed can be applied tomanipulation and detection of various types of analytes (includingbiomarkers). The analytes and are herein disclosed, or listed,described, and summarized in PCT Application (designating U.S.) Nos.PCT/US2016/045437 and PCT/US0216/051775, which were respectively filedon Aug. 10, 2016 and Sep. 14, 2016, U.S. Provisional Application No.62/456,065, which was filed on Feb. 7, 2017, U.S. ProvisionalApplication No. 62/456,287, which was filed on Feb. 8, 2017, and U.S.Provisional Application No. 62/456,504, which was filed on Feb. 8, 2017,all of which applications are incorporated herein in their entiretiesfor all purposes.

-   (8) Applications (Field and Samples)

The devices, systems, and methods herein disclosed can be used forvarious applications (fields and samples). The applications are hereindisclosed, or listed, described, and summarized in PCT Application(designating U.S.) Nos. PCT/US2016/045437 and PCT/US0216/051775, whichwere respectively filed on Aug. 10, 2016 and Sep. 14, 2016, U.S.Provisional Application No. 62/456,065, which was filed on Feb. 7, 2017,U.S. Provisional Application No. 62/456,287, which was filed on Feb. 8,2017, and U.S. Provisional Application No. 62/456,504, which was filedon Feb. 8, 2017, all of which applications are incorporated herein intheir entireties for all purposes.

-   (9) Cloud

The devices, systems, and methods herein disclosed can employ cloudtechnology for data transfer, storage, and/or analysis. The relatedcloud technologies are herein disclosed, or listed, described, andsummarized in PCT Application (designating U.S.) Nos. PCT/US2016/045437and PCT/US0216/051775, which were respectively filed on Aug. 10, 2016and Sep. 14, 2016, U.S. Provisional Application No. 62/456,065, whichwas filed on Feb. 7, 2017, U.S. Provisional Application No. 62/456,287,which was filed on Feb. 8, 2017, and U.S. Provisional Application No.62/456,504, which was filed on Feb. 8, 2017, all of which applicationsare incorporated herein in their entireties for all purposes.

-   Additional Notes

Further examples of inventive subject matter according to the presentdisclosure are described in the following enumerated paragraphs.

It must be noted that as used herein and in the appended claims, thesingular forms “a”, “an”, and “the” include plural referents unless thecontext clearly dictates otherwise, e.g., when the word “single” isused. For example, reference to “an analyte” includes a single analyteand multiple analytes, reference to “a capture agent” includes a singlecapture agent and multiple capture agents, reference to “a detectionagent” includes a single detection agent and multiple detection agents,and reference to “an agent” includes a single agent and multiple agents.

As used herein, the terms “adapted” and “configured” mean that theelement, component, or other subject matter is designed and/or intendedto perform a given function. Thus, the use of the terms “adapted” and“configured” should not be construed to mean that a given element,component, or other subject matter is simply “capable of” performing agiven function. Similarly, subject matter that is recited as beingconfigured to perform a particular function may additionally oralternatively be described as being operative to perform that function.

As used herein, the phrase, “for example,” the phrase, “as an example,”and/or simply the terms “example” and “exemplary” when used withreference to one or more components, features, details, structures,embodiments, and/or methods according to the present disclosure, areintended to convey that the described component, feature, detail,structure, embodiment, and/or method is an illustrative, non-exclusiveexample of components, features, details, structures, embodiments,and/or methods according to the present disclosure. Thus, the describedcomponent, feature, detail, structure, embodiment, and/or method is notintended to be limiting, required, or exclusive/exhaustive; and othercomponents, features, details, structures, embodiments, and/or methods,including structurally and/or functionally similar and/or equivalentcomponents, features, details, structures, embodiments, and/or methods,are also within the scope of the present disclosure.

As used herein, the phrases “at least one of” and “one or more of,” inreference to a list of more than one entity, means any one or more ofthe entity in the list of entity, and is not limited to at least one ofeach and every entity specifically listed within the list of entity. Forexample, “at least one of A and B” (or, equivalently, “at least one of Aor B,” or, equivalently, “at least one of A and/or B”) may refer to Aalone, B alone, or the combination of A and B.

As used herein, the term “and/or” placed between a first entity and asecond entity means one of (1) the first entity, (2) the second entity,and (3) the first entity and the second entity. Multiple entity listedwith “and/or” should be construed in the same manner, i.e., “one ormore” of the entity so conjoined. Other entity may optionally be presentother than the entity specifically identified by the “and/or” clause,whether related or unrelated to those entities specifically identified.

Where numerical ranges are mentioned herein, the invention includesembodiments in which the endpoints are included, embodiments in whichboth endpoints are excluded, and embodiments in which one endpoint isincluded and the other is excluded. It should be assumed that bothendpoints are included unless indicated otherwise. Furthermore, unlessotherwise indicated or otherwise evident from the context andunderstanding of one of ordinary skill in the art.

In the event that any patents, patent applications, or other referencesare incorporated by reference herein and (1) define a term in a mannerthat is inconsistent with and/or (2) are otherwise inconsistent with,either the non-incorporated portion of the present disclosure or any ofthe other incorporated references, the non-incorporated portion of thepresent disclosure shall control, and the term or incorporateddisclosure therein shall only control with respect to the reference inwhich the term is defined and/or the incorporated disclosure was presentoriginally.

What is claimed is:
 1. A device for assaying a sample using light,comprising: a first plate, a second plate, spacers, and a texturedsurface, wherein: (i) the plates are movable relative to each other intodifferent configurations; (ii) one or both plates are flexible; (iii)the first plate is transparent to the light; (iv) the second plate has,on its inner surface, a textured structure for scattering the lightilluminated on the surface; (v) the textured surface is a bumpy, wavyrough surface; and (vi) the spacers are fixed to the inner surface ofone of the plates, and have a predetermined uniform height that islarger than the average roughness of the textured surface but smallerthan 200 μm; wherein one of the configurations is an open configuration,in which: the two plates are partially or entirely separated apart, thespacing between the two plates is not regulated by the spacers, and thesample is deposited on one or both of the plates; wherein one of theconfigurations is a closed configuration, which is configured after thesample deposition in the open configuration, and in the closedconfiguration: at least part of the deposited sample is compressed bythe two plates into a continuous layer.
 2. An apparatus, comprising: a)the device of claim 1; b) a mobile computing device having a cameramodule; c) a light source, and d) an external lens.
 3. A method forassaying a sample using light, comprising the steps of: a) obtaining asample that contains or is suspected of containing an analyte; b)obtaining the device of claim 1; c) depositing the sample on one or bothof the plates of the device when the plates are in the openconfiguration; d) bringing the two plates together and pressing theplates into the closed configuration so that the sample forms a thinlayer between the two plates; and e) measuring, after step (d), anoptical signal from the sample.
 4. The device of claim 1, wherein thetextured surface has an average roughness range of 2 μm to 5 μm.
 5. Thedevice of claim 1, wherein the textured surface is made of semi-opaquewhite material, and the transmissivity is 10%˜30%.
 6. The device ofclaim 1, wherein the texture on the textured surface is periodic.
 7. Thedevice of claim 1, wherein the textured surface is made of semi-opaquewhite material, and has a transmissivity of 10% to 30%.
 8. The device ofclaim 1, wherein the textured surface is opaque white material or coatedwith reflective metal film, the metal film selected from aluminum,silver or gold.
 9. The device of claim 1, wherein the textured surfaceis made of a highly reflectively opaque white material with reflectivityof at least 50%, 60%, 70%, 80%, 90%, 100%, or in a range between any ofthe two values.
 10. The device of claim 1, wherein the reflectionspectrum of the textured structure is within the range of 300 nm to 1000nm.
 11. The device of claim 1, wherein the texture surface is made of asemi-opaque white material, and has a transmissivity of 10%˜30%.
 12. Thedevice of claim 1, wherein the texture surface is made of a reflectivemetal film.
 13. The device of claim 1, wherein the textured surface hasan arithmetic average roughness of 0.5 μm˜200 μm, and a mean spacing ofthe asperities of >0.5 μm and an average slop of the profile is greaterof 0.1.
 14. The device of claim 1, wherein the textured surface is madeof opaque white material.
 15. The device of claim 1, wherein the averagelateral feature size of the textured surface is at least 20% and up to10-fold of the wavelength of the light, or in range between these twovalues.
 16. The device of claim 1, wherein the average lateral featuresize of the textured surface is at least 50% and up to 1000-fold of thewavelength of the light, or in range between these two values.
 17. Thedevice of claim 1, wherein the textured surface has a surface roughnessat least 20% of the wavelength of the light.
 18. The device of claim 1,wherein the flexible plate has the thickness of the flexible plate timesthe Young's modulus of the flexible plate in the range 60 to 750 GPa-μm,and wherein the fourth power of the inter-spacer-distance (ISD) dividedby the thickness of the flexible plate (h) and the Young's modulus (E)of the flexible plate, ISD⁴/(hE), is equal to or less than 10⁶ μm³/GPa.19. The device of claim 1, wherein the spacer has a height of 30 μm orless.
 20. The device of claim 1, further comprising one or more reagentson one of the plates.
 21. The device of claim 1, wherein one or bothplates further comprise a location marker, a scale marker, an imagemarker, or any combination of thereof.
 22. The apparatus of claim 2,wherein the light source emits white light.
 23. The apparatus of claim2, wherein the distance between the plates and the camera module of themobile computing device is in a range of 15 mm to 20 mm.
 24. Theapparatus of claim 2, wherein the external lens is put between theplates and the camera module, so that the sample between the plates isin a working distance of the camera module, and a focal length of theexternal lens is 12 to 18 mm, and the distance between the external lensand the camera module is no larger than 3 mm.
 25. The apparatus of claim2, wherein the distance between the light source and the plates is in arange of 5 mm to 10 mm.
 26. The apparatus of claim 2, wherein thedistance between the plates and the camera module is 5 to 10 mm.
 27. Theapparatus of claim 2, wherein the distance between the external lens isput between the plates and the camera module, so that the sample betweenthe plates is in a working distance of the camera module, and a focallength of the external lens is 4 to 8 mm, and the distance between theexternal lens and the camera module is no larger than 3 mm.
 28. Theapparatus of claim 2, wherein the light source comprises an opticalfiber guide that emits light from side and forms a ring shape.
 29. Themethod of claim 3, wherein the optical signal is luminescence signal.30. The method of claim 3, wherein the optical signal is colorimetricsignal.
 31. The method of claim 3, wherein the analyte is selected fromthe group of pH, ammonia, nitrite, nitrate, heavy metal, bacteria,lactose, food additive, and protein.
 32. The method of claim 3, whereinthe analyte is selected from the group consisting of glucose, albumin,sodium, potassium, chloride, urea nitrogen, creatinine, alkalinephosphatase, alanine amino transferase, aspartate amino transferase,bilirubin, cholesterol, triglycerides, hydrogen peroxide, and alcohol.33. The method of claim 3, wherein the sample comprises a body fluidselected from the group consisting of: amniotic fluid, aqueous humour,vitreous humour, blood (e.g., whole blood, fractionated blood, plasma,serum, etc.), breast milk, cerebrospinal fluid (CSF), cerumen (earwax),chyle, chime, endolymph, perilymph, feces, gastric acid, gastric juice,lymph, mucus (including nasal drainage and phlegm), pericardial fluid,peritoneal fluid, pleural fluid, pus, rheum, saliva, sebum (skin oil),semen, sputum, sweat, synovial fluid, tears, vomit, urine and exhaledcondensate.
 34. The method of claim 3, wherein the sample comprises anenvironmental specimen that is obtained from: river, lake, pond, ocean,glaciers, icebergs, rain, snow, sewage, reservoirs, tap water, drinkingwater, soil, compost, sand, rocks, concrete, wood, brick, sewage; air,heat vents, industrial exhaust, or vehicular exhaust.
 35. The method ofclaim 3, wherein the sample comprises a foodstuff specimen thatincludes: raw food ingredients, cooked or processed food, plant andanimal sources of food, preprocessed food, or fully processed food.